Therapeutic compositions containing harmine and isovanillin components, and methods of use thereof

ABSTRACT

Human therapeutic treatment compositions comprise at least two of a curcumin component, a harmine component, and an isovanillin component, and preferably all three in combination. The agents are effective for the treatment of human conditions, especially human cancers.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. application Ser. No.15/826,101 filed Nov. 29, 2017, which is a continuation of U.S.application Ser. No. 15/337,987 filed Oct. 28, 2016, which is acontinuation-in-part of PCT application SN PCT/US2015/055968 filed Oct.16, 2015, which is a continuation-in-part of U.S. utility applicationSer. No. 14/721,011 filed May 26, 2015 (issued Aug. 2, 2016 as U.S. Pat.No. 9,402,834), and which claims the benefit of U.S. ProvisionalApplications, Ser. 62/184,051 filed Jun. 24, 2015, Ser. 62/161,090 filedMay 13, 2015, and Ser. 62/066,686 filed Oct. 21, 2014. U.S. applicationSer. No. 15/337,987 filed Oct. 28, 2016, is also a continuation of PCTApplication SN PCT/M2016/000723 filed Apr. 20, 2016, which is acontinuation-in-part of U.S. utility application Ser. No. 14/721,011filed May 26, 2015 (issued Aug. 2, 2016 as U.S. Pat. No. 9,402,834), andwhich claims the benefit of U.S. Provisional Application Ser. 62/184,051filed Jun. 24, 2015, and Ser. 62/161,090 filed May 13, 2015. Each of theabove provisional, non-provisional, and PCT applications is incorporatedby reference herein in its entirety.

BACKGROUND OF THE INVENTION Field of the Invention

The present invention relates to combination chemotherapeutics fortreatment of humans, and especially for the treatment of human cancers,and corresponding methods for the treatment of humans suffering fromcancers or other maladies. The invention further provides dosage formsand regimens for administration to human patients, and methods offormulating and administering such dosage forms to yield improvements intreatment outcomes. More particularly, the invention is concerned withthe administration of specific chemotherapeutic dosage forms (e.g.,liquid mixtures, capsules, pills, or tablets) containing one or morecurcumin component(s), harmine component(s), and isovanillincomponent(s), and sub-combinations thereof.

Description of Related Art

Cancer is a generic term for a large group of diseases that can affectany part of the body. Other terms used are malignant tumors andneoplasms. One defining feature of cancer is the rapid creation ofabnormal cells that grow beyond their usual boundaries, and which canthen invade adjoining parts of the body and spread to other organs. Thisprocess is referred to as metastasis.

Metastases are the major cause of death from cancer.

The transformation from a normal cell into a tumor cell is a multistageprocess, typically a progression from a pre-cancerous lesion tomalignant tumors. These changes are the result of the interactionbetween a person's genetic factors and three categories of externalagents, including:

-   -   physical carcinogens, such as ultraviolet and ionizing radiation    -   chemical carcinogens, such as asbestos, components of tobacco        smoke, aflatoxin (a food contaminant) and arsenic (a drinking        water contaminant)    -   biological carcinogens, such as infections from certain viruses,        bacteria or parasites.

Some examples of infections associated with certain cancers:

-   -   Viruses: hepatitis B and liver cancer, Human Papilloma Virus        (HPV) and cervical cancer, and human immunodeficiency virus        (HIV) and Kaposi sarcoma.    -   Bacteria: Helicobacter pylori and stomach cancer.    -   Parasites: schistosomiasis and bladder cancer.

Aging is another fundamental factor for the development of cancer. Theincidence of cancer rises dramatically with age, most likely due to abuildup of risks for specific cancers that increase with age. Theoverall risk accumulation is combined with the tendency for cellularrepair mechanisms to be less effective as a person grows older.

Tobacco use, alcohol use, low fruit and vegetable intake, and chronicinfections from hepatitis B (HBV), hepatitis C virus (HCV) and sometypes of Human Papilloma Virus (HPV) are leading risk factors for cancerin low- and middle-income countries. Cervical cancer, which is caused byHPV, is a leading cause of cancer death among women in low-incomecountries. In high-income countries, tobacco use, alcohol use, and beingoverweight or obese are major risk factors for cancer.

The most common cancer treatment modalities are surgery, chemotherapy,and radiation treatments. All of these techniques have significantdrawbacks in terms of side effects and patient discomfort. For example,chemotherapy may result in significant decreases in white blood cellcount (neutropenia), red blood cell count (anemia), and platelet count(thrombocytopenia). This can result in pain, diarrhea, constipation,mouth sores, hair loss, nausea, and vomiting.

Biological therapy (sometimes called immunotherapy, biotherapy, orbiological response modifier therapy) is a relatively new addition tothe family of cancer treatments. Biological therapies use the body'simmune system, either directly or indirectly, to fight cancer or tolessen the side effects that may be caused by some cancer treatments.

During chemotherapies involving multiple-drug treatments, adverse drugevents are common, and indeed toxicities related to drug-druginteractions are one of the leading causes of hospitalizations in theUS. Obach, R. S. “Drug-Drug Interactions: An Important NegativeAttribute in Drugs.” Drugs Today 39.5 (2003): 308-338. In fact, in anysingle-month period, one-fifth of all surveyed adults in the USAreported an adverse drug response. Hakkarainen, K. M. et al. “Prevalenceand Perceived Preventability of Self-Reported Adverse Drug Events—APopulation-Based Survey of 7,099 Adults.” PLoS One 8.9 (2013): e73166. Alarge-scale study of adults aged 57-85 found that 29% were taking morethan five prescription medications and nearly 5% were at risk of majoradverse drug-drug interactions. In the field of oncology, a review ofover 400 cancer patients determined that 77% were taking drugs that wereconsidered to have a moderately severe potential for adverse druginteractions, and 9% had major adverse drug interactions. Ghalib, M. S.et al. “Alterations of Chemotherapeutic Pharmocokinetic Profiles byDrug-Drug Interactions.” Expert Opin. Drug Metabl. Toxicol 5.2 (2009):109-130.

Such interactions are a global health problem, and the WHO hasdetermined that negative drug interactions are leading causes ofmorbidity and mortality around the world, with up to 7% of allhospitalizations in the US due to negative drug interactions. A recentsurvey of a single hospital shows that 83% of hospitalized patients wereprescribed drug combinations with the potential to cause adversereactions. Patel, P. S. et al. “A Study of Potential Adverse Drug-DrugInteractions Among Prescribed Drugs in a Medicine Outpatient Departmentof a Tertiary Care Teaching Hospital.” J. Basic Clin. Pharm. 5.2 (2014):44-48.

Examples of famous negative drug interactions include the development ofrhabdomyolysis, a severe muscle disease, when taking Simvastatin withAmiodarone. As a result, the FDA introduced a warning on the drug labelabout the interaction. The calcium channel blocker Mibefradif, taken forhigh blood pressure, was removed from the market because of the harmfulinteraction with drugs that work on the electrical activity of theheart.

Cancer cells are cells that, by definition, grow and divide withoutnormal limitations. The unrestricted cell growth results in tumors,comprised of a variety of cell types. Treatments to fight cancer arefrequently successful in killing the typical, differentiated cancercells that form the majority of a solid tumor, otherwise known as thebulk cells. However even with the best treatment, the cancer may returna few months to years later (Prince, M. E. et al., “Cancer stem cells inhead and neck squamous cell cancer.” J. Clin. Oncol. 26.17(2008):2871-2875). For example, recurrence is frequently the case forpancreatic and head and neck cancer. It is now hypothesized that one ofthe key factors in the recurrence rate for cancers is the presence ofcancer stem cells.

Cancer stem cells were not identified until the late 1990s and show twoimportant properties of stem cells: 1) cancer stem cells can self-renewand, 2) cancer stem cells can differentiate into any other cell type(Bandhavkar, S. “Cancer stem cells: a metastasizing menace.” Cancer Med.(2016) doi:10.1002/cam4.629; Dick, J. E. “Stem cell concepts renewcancer research.” Blood. 112 (2008):4793-4807). While they make up onlya small percentage of the total number of cells in a tumor, theycompromise a unique category of cancer cells that are more likely to beresistant to chemotherapy or radiation therapy. In fact, it is nowbelieved that the majority of cells in tumors are not cancer-causing andcannot initiate new tumors (Bandhavkar). Only cancer stem cells appearto be tumor-initiators (Visvader, J. E. et al. “Cancer stem cells:Current status and evolving complexities.” Cell Stem Cell. 10(2012):717-728). Cancer stem cells have been shown to coordinate tumorcell growth, metastases (migration and invasion), and drug resistance(Cammarota, F. et al. “Mesenchymal stem/stromal cells in stromalevolution and cancer progression.” Stem Cells Int. (2016):4824573).These cancer stem cells behave differently than non-cancerous stem cellsin the person (Cammarota et al.), and have been described as the “rootsof aggressive tumors for which we have no effective treatment” (Doherty,M. R. et al. “Cancer stem cell plasticity drives therapeuticresistance.” Cancers 8,8 (2016) doi:10.3390). In general stem cells arenaturally resistant to chemotherapies and radiation therapy (Diehn, M.et al. “Cancer stem cells and radiotherapy: new insights into tumorradioresistance.” J. Natl. Cancer Inst. 98 (2016):1775-1757; Mery, B. etal. “Targeting head and neck tumoral stem cells: From biological aspectsto therapeutic perspectives.” World J Stem Cells 8.1 (2016):13-21),because they have chemical pumps that remove the chemotherapies out ofthe cells, thus they can have no effect on the stem cells. They alsohave a slow rate of turnover, and most radiation and chemotherapies aredesigned to only kill cells that are rapidly dividing, such as themajority of the cells within the tumor. These characteristics explain,on a large scale, how stem cells associated with cancer are resistant tochemotherapy.

Current anticancer therapies may directly cause cancer cells to die orjust inhibit their growth (Bandhavkar). If the anticancer therapy failsto target and remove the cancer stem cells, then relapse and drugresistance ensues. Selectively targeting and eliminating cancer stemcells would theoretically treat the primary tumors and halt any chanceof recurrence (Mery et al.). Yet, the ability to kill cancer stem cellsis currently considered a significant clinical challenge. Some recentevidence suggests that traditional chemotherapies can even induce thegeneration of new stem cells within tumors potentially making the cancerreturn faster (Doherty et. al).

Identification of the regulatory mechanisms and signaling pathwaysinvolved in cancer stem cells (CSCs) will help in designing novel agentsto target this refractory cell population in pancreatic cancers. Cancerstem cells are capable of self-renewal and generating tumors resemblingthe primary tumor (Ponnurangam, S. et al. “Quinomycin A targets Notchsignaling pathway in pancreatic cancer stem cells.” Oncotarget 7.3(2015):3217-3232). The sphere-forming assays have been widely used toidentify stem cells based on their reported capacity to evaluateself-renewal and differentiation.

U.S. Pat. No. 8,039,025 describes cancer treatments in the form ofextracts of Arum palaestinum Boiss, supplemented with individual amountsof β-sitosterol, isovanillin, and linoleic acid, and this patent isincorporated by reference herein in its entirety.

Despite the immense amount of worldwide research and efforts to stem thetide of cancer and its side effects, the disease in its manymanifestations continues to be a huge problem. Therefore, any new cancertreatment having a curative affect and/or the ability to amelioratecancer symptoms and improve the lifestyle of patients is highlysignificant and important.

SUMMARY OF THE INVENTION

The present invention provides improved chemotherapeutics for treatmentof humans, and especially in the treatment of human cancers, with novelcombinations of compounds, which are useful against a wide variety ofdifferent cancers with minimal or nonexistent adverse side effects.Generally speaking, the chemotherapeutics of the invention compriserespective quantities of at least two of, curcumin component(s), harminecomponent(s), and isovanillin component(s). The preferredchemotherapeutics include all three of these components, butsub-combinations thereof are also useful, i.e., therapeutics comprisingcurcumin component(s) and harmine component(s), curcumin component(s)and isovanillin component(s), and harmine component(s) and isovanillincomponent(s). In particularly preferred embodiments, the activecomponents of the compositions (i.e., those having a significanttherapeutic effect) consist essentially of curcumin, harmine, andisovanillin components in the case of three-component compositions, andconsist essentially of two of the three components in the case of thetwo-component compositions. Preferably, the at least one curcumincomponent comprises curcumin, the at least one harmine componentcomprises harmine, and the at least one isovanillin component comprisesisovanillin or vanillin.

The invention also provides new methods for treatment of cancers byadministration of appropriate quantities of the anti-cancer compositionshereof. Hence, the compositions are particularly designed for use in thetreatment of cancers, and the compositions can be used for themanufacture of medicaments of anti-cancer therapeutic applications. Inaddition, the invention provides pharmaceutical compositions for thetreatment of cancers comprising administering therapeutically effectiveamounts of the new compositions, prepared by processes known per se,with a pharmaceutically acceptable carrier.

Curcumin (CAS #458-37-7) is a diaryl heptanoid, and has the molecularformula C21H20O6. Curcumin occurs as a part of a curcuminoid plantextract containing curcumin, demethoxycurcumin, andbis-demethoxycurcumin. One commercially available effective curcuminoidis sold as “Curcumin from Curcuma Longa (Turmeric),” which containsgreater than 65% by weight curcumin, as determined by HPLC analysis.

Harmine (CAS #442-51-3) is a methoxy methyl pyrido indole belonging tothe β-carboline family of compounds, and has the molecular formulaC13H12N2O. Harmine occurs in a number of different plants native to theMiddle East and South America.

Isovanillin (CAS #621-59-0) is a phenolic aldehyde vanillin isomer, andhas the molecular formula C8H803.

A “chemotherapeutic” or “chemotherapeutic agent” as used herein refersto the combinations of chemical compounds described herein as useful inthe treatment of human conditions, especially human cancers.Chemotherapeutics may be cytostatic, selectively toxic or destructive ofcancerous tissue and/or cells, but also include indiscriminatelycytotoxic compounds used in cancer treatments.

The combination therapeutic agents of the invention have been found tobe effective in the treatment of a broad spectrum of human cancers, andalso to other conditions, such as elevated PSA counts in men. The broadscope of utility with the agents of the invention is in itself highlyunusual. However, this feature, together with the nonexistent or minimalside effects induced by the agents, represents a startling developmentin the art.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a graph of cell number versus dosage amounts of GZ17-6.02,illustrating the effect thereof in inducing the death of two differenttypes of human head and neck cancer cells, as described in Example 1;

FIG. 2A is a graph of cell number versus dosage amounts of GZ17-6.02,illustrating the effect thereof GZ17-6.02 in inducing the death ofpediatric leukemia cells, as described in Example 2;

FIG. 2B is a graph of cell number versus dosage amounts of GZ17-6.02,illustrating the effect thereof GZ17-6.02 in inducing the death ofpediatric osteosarcoma cells, as described in Example 2;

FIG. 3A is a graph of cell number versus dosage amounts of GZ17-6.02,illustrating the effect thereof GZ17-6.02 in inducing the death oflymphoma cells, as described in Example 3;

FIG. 3B is a graph of cell number versus dosage amounts of GZ17-6.02,illustrating the effect thereof GZ17-6.02 in inducing the death of lungcancer cells, as described in Example 3;

FIG. 4A is a graph of cell number versus dosage amounts of GZ17-6.02,illustrating the effect thereof GZ17-6.02 in inducing the death ofovarian cancer cells, as described in Example 4;

FIG. 4B is a graph of cell number versus dosage amounts of GZ17-6.02,illustrating the effect thereof in inducing the death of prostate cancercells, as described in Example 4;

FIG. 5A is a graph of cell number versus dosage amounts of GZ17-6.02,illustrating the effect thereof in inducing the death of human breastcancer cells, as described in Example 5;

FIG. 5B is a graph of cell number versus dosage amounts of GZ17-6.02,illustrating the effect thereof in inducing the death of pancreaticcancer cells, as described in Example 5;

FIG. 6 is a graph of cell number versus doses of GZ17-6.02 illustratingthe relative effects thereof in inducing cell death in prostate cancerand ovarian cancer cells, as compared with non-cancerous fibroblasts, asdescribed in Example 6;

FIG. 7A is a graph illustrating the effect of GZ17-6.02 in preventingmigration of head and neck cancer cells, as described in Example 7;

FIG. 7B is a graph illustrating the effect of GZ17-6.02 in preventinginvasion of head and neck cancer cells, as described in Example 7;

FIG. 8 is a graph of normalized cell number versus increasingdoxorubicin doses alone and with the addition of two differentconcentrations of GZ17-6.02, as described in Example 8;

FIG. 9A is a graph illustrating extent of apoptosis in a control test,as described in Example 9, and illustrating about 19.8% cell death viaapoptosis;

FIG. 9B is a graph illustrating extent of apoptosis in a test identicalto the control test of FIG. 9A, but including GZ17-6.02, as described inExample 9, and illustrating about 48.2% cell death via apoptosis;

FIG. 10A is a graph illustrating that caspase 3 and 7 concentrationsincreased in response to GZ17-6.02, but caspase 9 levels did not changein response to GZ17-6.02 in lung cancer cells, as described in Example10;

FIG. 10B is a graph illustrating that caspase 6 concentrations increasedin response to GZ17-6.02 in lung cancer cells, as described in Example10;

FIG. 10C is a graph illustrating that ATP levels, as a marker ofmitochondrial toxicity, were not increased by GZ17-6.02 in lung cancercells, as described in Example 10;

FIG. 11A is a graph illustrating the mechanisms of ovarian cancer cellsdeath by application of GZ17-6.02, as explained in Example 11;

FIG. 11B is a graph illustrating the mechanisms of ovarian cancer cellsdeath by application of GZ17-6.02, as explained in Example 11;

FIG. 11C is a graph illustrating the mechanisms of ovarian cancer cellsdeath by application of GZ17-6.02, as explained in Example 11;

FIG. 11D is a graph illustrating the mechanisms of ovarian cancer cellsdeath by application of GZ17-6.02, as explained in Example 11;

FIG. 11E is a graph illustrating the mechanisms of ovarian cancer cellsdeath by application of GZ17-6.02, as explained in Example 11;

FIG. 12A is a graph illustrating the mechanisms of osteosarcoma cellsdeath by application of GZ17-6.02, as explained in Example 12;

FIG. 12B is a graph illustrating the mechanisms of osteosarcoma cellsdeath by application of GZ17-6.02, as explained in Example 12;

FIG. 12C is a graph illustrating the mechanisms of osteosarcoma cellsdeath by application of GZ17-6.02, as explained in Example 12;

FIG. 12D is a graph illustrating the mechanisms of osteosarcoma cellsdeath by application of GZ17-6.02, as explained in Example 12;

FIG. 13 is a graph illustrating the mechanisms of human head and neckcancer cells death by application of GZ17-6.02, as explained in Example13;

FIG. 14A is a control quantitative dot-blot, which measures the relativeamount of proteins, known to be involved in cell proliferation describedin Example 14, where the proteins were epidermal growth factor receptor(EGFR), extracellular-signal-regulated kinase (ERK1/2), the catalyticsubunit of AMP-activated kinase (AMPKα2), β-catenin, and Chk-2.

FIG. 14B is a quantitative dot-block similar to that described in FIG.14A, but illustrating the amounts of the test proteins upon applicationof GZ17-6.02, as described in Example 14;

FIG. 15A is a graph depicting the results of two independent scientists,each carrying out an identical induced cell death test with GZ17-6.02 onovarian cancer cells, as described in Example 15;

FIG. 15B is a graph depicting the results of two independent scientists,each carrying out an identical induced cell death test with GZ17-6.02 onlung cancer cells, as described in Example 4; and

FIG. 16A is a comparative graph of tumor volume versus days after cancercell inoculation in mice, between control inoculations (ethanol vehicle)and test inoculations containing the vehicle and GZ17-6.02, illustratingthe dramatic reduction in tumor volumes in the test mice, as explainedin Example 16;

FIG. 16B is a comparative graph of contralateral tumor volume versusdays after cancer cell inoculation in mice, indicating a systemic effectof use of GZ17-6.02, as explained in Example 16;

FIG. 16C is a graph of fractional tumor volume versus days aftertreatment wherein one group of mice was implanted with human head andneck tumor cells using an implantation vehicle, and a control group ofmice was implanted with only the vehicle, in order to determine tumorvolume over time, as set forth in Example 16;

FIG. 17A is a graph of cell number versus dosage amounts of GZ17-6.02(open circles) versus a combined product including ⅓ by weight of eachGZ17-6.02 component (filled circles), tested on ovarian cancer cells, asdescribed in Example 4;

FIG. 17B is a graph of cell number versus dosage amounts of GZ17-6.02(open circles) versus a combined product including ⅓ by weight of eachGZ17-6.02 component (filled circles), tested on lung cancer cells, asdescribed in Example 3;

FIG. 17C is a graph of cell number versus dosage amounts of GZ17-6.02(open circles) versus a combined product including ⅓ by weight of eachGZ17-6.02 component (filled circles), tested on prostate cancer cells,as described in Example 4;

FIG. 18A is a graph of percent ovarian cancer cell death versusdifferent component combinations of GZ17-6.02, illustrating resultsusing isovanillin alone, and two-component products respectivelyincluding isovanillin plus curcumin, and isovanillin plus harmine, wherethe isovanillin concentration was held constant throughout;

FIG. 18B is a graph of percent lung cancer cell death versus differentcomponent combinations of GZ17-6.02, illustrating results usingisovanillin alone, and two-component products respectively includingisovanillin plus curcumin, and isovanillin plus harmine, where theisovanillin concentration was held constant throughout;

FIG. 18C is a graph of percent prostate cancer cell death versusdifferent component combinations of GZ17-6.02, illustrating resultsusing isovanillin alone, and two-component products respectivelyincluding isovanillin plus curcumin, and isovanillin plus harmine, wherethe isovanillin concentration was held constant throughout;

FIG. 18D is a graph of percent lymphoma cancer cell death versusdifferent component combinations of GZ17-6.02, illustrating resultsusing isovanillin alone, and two-component products respectivelyincluding isovanillin plus curcumin, and isovanillin plus harmine, wherethe isovanillin concentration was held constant throughout;

FIG. 19A is a graph of percent ovarian cancer cell death versusdifferent component combinations of GZ17-6.02, illustrating resultsusing curcumin alone, and two-component products respectively includingcurcumin plus isovanillin, and curcumin plus harmine, where the curcuminconcentration was held constant throughout;

FIG. 19B is a graph of percent lung cancer cell death versus differentcomponent combinations of GZ17-6.02, illustrating results using curcuminalone, and two-component products respectively including curcumin plusisovanillin, and curcumin plus harmine, where the curcumin concentrationwas held constant throughout;

FIG. 19C is a graph of percent prostate cancer cell death versusdifferent component combinations of GZ17-6.02, illustrating resultsusing curcumin alone, and two-component products respectively includingcurcumin plus isovanillin, and curcumin plus harmine, where the curcuminconcentration was held constant throughout;

FIG. 19D is a graph of percent lymphoma cancer cell death versusdifferent component combinations of GZ17-6.02, illustrating resultsusing curcumin alone, and two-component products respectively includingcurcumin plus isovanillin, and curcumin plus harmine, where the curcuminconcentration was held constant throughout;

FIG. 20A is a graph of percent ovarian cancer cell death versusdifferent component combinations of GZ17-6.02, illustrating resultsusing harmine alone, and two-component products respectively includingharmine plus isovanillin, and harmine plus curcumin, where the harmineconcentration was held constant throughout;

FIG. 20B is a graph of percent lung cancer cell death versus differentcomponent combinations of GZ17-6.02, illustrating results using harminealone, and two-component products respectively including harmine plusisovanillin, and harmine plus curcumin, where the harmine concentrationwas held constant throughout;

FIG. 20C is a graph of percent prostate cancer cell death versusdifferent component combinations of GZ17-6.02, illustrating resultsusing harmine alone, and two-component products respectively includingharmine plus isovanillin, and harmine plus curcumin, where the harmineconcentration was held constant throughout;

FIG. 20D is a graph of percent lymphoma cancer cell death versusdifferent component combinations of GZ17-6.02, illustrating resultsusing harmine alone, and two-component products respectively includingharmine plus isovanillin, and harmine plus curcumin, where the harmineconcentration was held constant throughout;

FIG. 21A is a graph of lymphoma cancer cell lethality using GZ17-6.02 ata dosage rate of 12 μg/mL, and using the three components of GZ17-6.02individually at the concentration present in GZ17-6.02, and furtherillustrating the theoretical additive effect of the three componentsversus GZ17-6.02;

FIG. 21B is a graph of lymphoma cancer cell lethality using GZ17-6.02 ata dosage rate of 24 μg/mL, and using the three components of GZ17-6.02individually at the concentration present in GZ17-6.02, and furtherillustrating the theoretical additive effect of the three componentsversus GZ17-6.02;

FIG. 21C is a graph of lymphoma cancer cell lethality using GZ17-6.02 ata dosage rate of 48 μg/mL, and using the three components of GZ17-6.02individually at the concentration present in GZ17-6.02, and furtherillustrating the theoretical additive effect of the three componentsversus GZ17-6.02;

FIG. 21D is a graph of lymphoma cancer cell lethality using GZ17-6.02 ata dosage rate of 96 μg/mL, and using the three components of GZ17-6.02individually at the concentration present in GZ17-6.02, and furtherillustrating the theoretical additive effect of the three componentsversus GZ17-6.02;

FIG. 22A is a graph of ovarian cancer cell lethality using GZ17-6.02 ata dosage rate of 12 μg/mL, and using the three components of GZ17-6.02individually at the concentration present in GZ17-6.02, and furtherillustrating the theoretical additive effect of the three componentsversus GZ17-6.02;

FIG. 22B is a graph of ovarian cancer cell lethality using GZ17-6.02 ata dosage rate of 24 μg/mL, and using the three components of GZ17-6.02individually at the concentration present in GZ17-6.02, and furtherillustrating the theoretical additive effect of the three componentsversus GZ17-6.02;

FIG. 22C is a graph of breast cancer cell lethality using GZ17-6.02 at adosage rate of 24 μg/mL, and using the three components of GZ17-6.02individually at the concentration present in GZ17-6.02, and furtherillustrating the theoretical additive effect of the three componentsversus GZ17-6.02;

FIG. 23A is a graph of lung cancer cell number versus increasing dosageamounts of vanillin alone, as described in Example 1;

FIG. 23B is a graph of lung cancer cell number versus increasing dosageamounts of isovanillic acid alone, as described in Example 1;

FIG. 23C is a graph of lung cancer cell number versus increasing dosageamounts of O-vanillin alone, as described in Example 1;

FIG. 23D is a graph of lung cancer cell number versus increasing dosageamounts of isovanillyl alcohol alone, as described in Example 1;

FIG. 23E is a graph of ovarian cancer cell number versus increasingdosage amounts of vanillin alone, as described in Example 1;

FIG. 23F is a graph of ovarian cancer cell number versus increasingdosage amounts of isovanillic acid alone, as described in Example 1;

FIG. 23G is a graph of ovarian cancer cell number versus increasingdosage amounts of O-vanillin alone, as described in Example 1;

FIG. 23H is a graph of ovarian cancer cell number versus increasingdosage amounts of isovanillyl alcohol alone, as described in Example 1;

FIG. 23I is a graph of prostate cancer cell number versus increasingdosage amounts of vanillin alone, as described in Example 1;

FIG. 23J is a graph of prostate cancer cell number versus increasingdosage amounts of isovanillic acid alone, as described in Example 1;

FIG. 23K is a graph of prostate cancer cell number versus increasingdosage amounts of O-vanillin alone, as described in Example 1;

FIG. 23L is a graph of prostate cancer cell number versus increasingdosage amounts of isovanillyl alcohol alone, as described in Example 1;

FIG. 24A is a graph of lung cancer cell number versus increasing dosageamounts of harmaline alone, as described in Example 1;

FIG. 24B is a graph of lung cancer cell number versus increasing dosageamounts of tetrahydro-harmine alone, as described in Example 1;

FIG. 24C is a graph of lung cancer cell number versus increasing dosageamounts of harmol hydrochloride alone, as described in Example 1;

FIG. 24D is a graph of lung cancer cell number versus increasing dosageamounts of harmalol hydrochloride dihydrate alone, as described inExample 1;

FIG. 24E is a graph of lung cancer cell number versus increasing dosageamounts of harmane alone, as described in Example 1;

FIG. 24F is a graph of ovarian cancer cell number versus increasingdosage amounts of harmaline alone, as described in Example 1;

FIG. 24G is a graph of ovarian cancer cell number versus increasingdosage amounts of tetrahydro-harmine alone, as described in Example 1;

FIG. 24H is a graph of ovarian cancer cell number versus increasingdosage amounts of harmol hydrochloride alone, as described in Example 1;

FIG. 24I is a graph of ovarian cancer cell number versus increasingdosage amounts of harmalol hydrochloride dihydrate alone, as describedin Example 1;

FIG. 24J is a graph of ovarian cancer cell number versus increasingdosage amounts of harmane alone, as described in Example 1;

FIG. 24K is a graph of prostate cancer cell number versus increasingdosage amounts of harmaline alone, as described in Example 1;

FIG. 24L is a graph of prostate cancer cell number versus increasingdosage amounts of tetrahydro-harmine alone, as described in Example 1;

FIG. 24M is a graph of prostate cancer cell number versus increasingdosage amounts of harmol hydrochloride alone, as described in Example 1;

FIG. 24N is a graph of prostate cancer cell number versus increasingdosage amounts of harmalol hydrochloride dihydrate alone, as describedin Example 1;

FIG. 24O is a graph of prostate cancer cell number versus increasingdosage amounts of harmane alone, as described in Example 1;

FIG. 25A is a graph of lung cancer cell number versus increasing dosageamounts of bisdemethoxy curcumin alone, as described in Example 1;

FIG. 25B is a graph of ovarian cancer cell number versus increasingdosage amounts of bisdemethoxy curcumin alone, as described in Example1;

FIG. 25C is a graph of prostate cancer cell number versus increasingdosage amounts of bisdemethoxy curcumin alone, as described in Example1;

FIG. 26 is a graph of ovarian cancer cell number versus dosage amountsof GZ17-6.02 where the product was stored at varying temperatures overtwo months, confirming that the product has long-term stability;

FIG. 27 is a graph of ovarian cancer cell number versus dosage amountsof GZ17-6.02, where the GZ17-6.02 was subjected to a series ofsuccessive freeze/thaw cycles, confirming that the product has excellentfreeze/thaw stability;

FIG. 28A is a graph of cell number versus dosage amounts of GZ17-8.02,illustrating the effect thereof in inducing the death of two differenttypes ovarian cancer, lung cancer, prostate cancer, pancreatic cancerand fibroblast cells, as described in Example 28;

FIG. 28B is a graph of cell number versus dosage amounts of GZ17-8.03,illustrating the effect thereof in inducing the death of ovarian cancer,lung cancer, prostate cancer, pancreatic cancer and fibroblast cells, asdescribed in Example 28;

FIG. 28C is a graph of cell number versus dosage amounts of GZ17-8.04,illustrating the effect thereof in inducing the death of ovarian cancer,lung cancer, prostate cancer, pancreatic cancer and fibroblast cells, asdescribed in Example 28;

FIG. 28D is a graph of cell number versus dosage amounts of GZ17-8.05,illustrating the effect thereof in inducing the death of ovarian cancer,lung cancer, prostate cancer, pancreatic cancer and fibroblast cells, asdescribed in Example 28;

FIG. 28E is a graph of cell number versus dosage amounts of GZ17-8.06,illustrating the effect thereof in inducing the death of ovarian cancer,lung cancer, prostate cancer, pancreatic cancer and fibroblast cells, asdescribed in Example 28;

FIG. 28F is a graph of cell number versus dosage amounts of GZ17-8.07,illustrating the effect thereof in inducing the death of ovarian cancer,lung cancer, prostate cancer, pancreatic cancer and fibroblast cells, asdescribed in Example 28;

FIG. 28G is a graph of cell number versus dosage amounts of GZ17-8.08,illustrating the effect thereof in inducing the death of ovarian cancer,lung cancer, prostate cancer, pancreatic cancer and fibroblast cells, asdescribed in Example 28;

FIG. 28H is a graph of cell number versus dosage amounts of GZ17-8.09,illustrating the effect thereof in inducing the death of ovarian cancer,lung cancer, prostate cancer, pancreatic cancer and fibroblast cells, asdescribed in Example 28;

FIG. 28I is a graph of cell number versus dosage amounts of GZ17-8.10,illustrating the effect thereof in inducing the death of ovarian cancer,lung cancer, prostate cancer, pancreatic cancer and fibroblast cells, asdescribed in Example 28;

FIG. 29A is a graph of cell number versus dosage amounts of GZ17-8.11,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 29B is a graph of cell number versus dosage amounts of GZ17-8.11,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 29C is a graph of cell number versus dosage amounts of GZ17-8.11,illustrating the effect thereof in inducing the death of prostatecancer;

FIG. 29D is a graph of cell number versus dosage amounts of GZ17-8.11,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 29E is a graph of cell number versus dosage amounts of GZ17-8.11,illustrating the effect thereof in inducing the death of breast cancer;

FIG. 29F is a graph of cell number versus dosage amounts of GZ17-8.11,illustrating the effect thereof in inducing the death of leukemia;

FIG. 29G is a graph of cell number versus dosage amounts of GZ17-8.11,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 30A is a graph of cell number versus dosage amounts of GZ17-8.12,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 30B is a graph of cell number versus dosage amounts of GZ17-8.12,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 30C is a graph of cell number versus dosage amounts of GZ17-8.12,illustrating the effect thereof in inducing the death of prostatecancer;

FIG. 30D is a graph of cell number versus dosage amounts of GZ17-8.12,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 30E is a graph of cell number versus dosage amounts of GZ17-8.12,illustrating the effect thereof in inducing the death of breast cancer;

FIG. 30F is a graph of cell number versus dosage amounts of GZ17-8.12,illustrating the effect thereof in inducing the death of leukemia;

FIG. 30G is a graph of cell number versus dosage amounts of GZ17-8.12,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 31A is a graph of cell number versus dosage amounts of GZ17-8.13,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 31B is a graph of cell number versus dosage amounts of GZ17-8.13,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 31C is a graph of cell number versus dosage amounts of GZ17-8.13,illustrating the effect thereof in inducing the death of prostatecancer;

FIG. 31D is a graph of cell number versus dosage amounts of GZ17-8.13,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 31E is a graph of cell number versus dosage amounts of GZ17-8.13,illustrating the effect thereof in inducing the death of breast cancer;

FIG. 31F is a graph of cell number versus dosage amounts of GZ17-8.13,illustrating the effect thereof in inducing the death of leukemia;

FIG. 31G is a graph of cell number versus dosage amounts of GZ17-8.13,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 32A is a graph of cell number versus dosage amounts of GZ17-8.14,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 32B is a graph of cell number versus dosage amounts of GZ17-8.14,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 32C is a graph of cell number versus dosage amounts of GZ17-8.14,illustrating the effect thereof in inducing the death of prostatecancer;

FIG. 32D is a graph of cell number versus dosage amounts of GZ17-8.14,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 32E is a graph of cell number versus dosage amounts of GZ17-8.14,illustrating the effect thereof in inducing the death of breast cancer;

FIG. 32F is a graph of cell number versus dosage amounts of GZ17-8.14,illustrating the effect thereof in inducing the death of leukemia;

FIG. 32G is a graph of cell number versus dosage amounts of GZ17-8.14,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 33A is a graph of cell number versus dosage amounts of GZ17-8.15,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 33B is a graph of cell number versus dosage amounts of GZ17-8.15,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 33C is a graph of cell number versus dosage amounts of GZ17-8.15,illustrating the effect thereof in inducing the death of prostatecancer;

FIG. 33D is a graph of cell number versus dosage amounts of GZ17-8.15,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 33E is a graph of cell number versus dosage amounts of GZ17-8.15,illustrating the effect thereof in inducing the death of breast cancer;

FIG. 33F is a graph of cell number versus dosage amounts of GZ17-8.15,illustrating the effect thereof in inducing the death of leukemia;

FIG. 33G is a graph of cell number versus dosage amounts of GZ17-8.15,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 34A is a graph of cell number versus dosage amounts of GZ17-8.16,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 34B is a graph of cell number versus dosage amounts of GZ17-8.16,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 34C is a graph of cell number versus dosage amounts of GZ17-8.16,illustrating the effect thereof in inducing the death of prostatecancer;

FIG. 34D is a graph of cell number versus dosage amounts of GZ17-8.16,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 34E is a graph of cell number versus dosage amounts of GZ17-8.16,illustrating the effect thereof in inducing the death of breast cancer;

FIG. 34F is a graph of cell number versus dosage amounts of GZ17-8.16,illustrating the effect thereof in inducing the death of leukemia;

FIG. 34G is a graph of cell number versus dosage amounts of GZ17-8.16,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 35A is a graph of cell number versus dosage amounts of GZ17-8.17,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 35B is a graph of cell number versus dosage amounts of GZ17-8.17,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 35C is a graph of cell number versus dosage amounts of GZ17-8.17,illustrating the effect thereof in inducing the death of prostatecancer;

FIG. 35D is a graph of cell number versus dosage amounts of GZ17-8.17,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 35E is a graph of cell number versus dosage amounts of GZ17-8.17,illustrating the effect thereof in inducing the death of breast cancer;

FIG. 35F is a graph of cell number versus dosage amounts of GZ17-8.17,illustrating the effect thereof in inducing the death of leukemia;

FIG. 35G is a graph of cell number versus dosage amounts of GZ17-8.17,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 36A is a graph of cell number versus dosage amounts of GZ17-8.18,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 36B is a graph of cell number versus dosage amounts of GZ17-8.18,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 36C is a graph of cell number versus dosage amounts of GZ17-8.18,illustrating the effect thereof in inducing the death of prostatecancer;

FIG. 36D is a graph of cell number versus dosage amounts of GZ17-8.18,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 36E is a graph of cell number versus dosage amounts of GZ17-8.18,illustrating the effect thereof in inducing the death of breast cancer;

FIG. 36F is a graph of cell number versus dosage amounts of GZ17-8.18,illustrating the effect thereof in inducing the death of leukemia;

FIG. 36G is a graph of cell number versus dosage amounts of GZ17-8.18,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 37A is a graph of cell number versus dosage amounts of GZ17-8.19,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 37B is a graph of cell number versus dosage amounts of GZ17-8.19,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 37C is a graph of cell number versus dosage amounts of GZ17-8.19,illustrating the effect thereof in inducing the death of prostatecancer;

FIG. 37D is a graph of cell number versus dosage amounts of GZ17-8.19,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 37E is a graph of cell number versus dosage amounts of GZ17-8.19,illustrating the effect thereof in inducing the death of breast cancer;

FIG. 37F is a graph of cell number versus dosage amounts of GZ17-8.19,illustrating the effect thereof in inducing the death of leukemia;

FIG. 37G is a graph of cell number versus dosage amounts of GZ17-8.19,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 38A is a graph of cell number versus dosage amounts of GZ17-8.20,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 38B is a graph of cell number versus dosage amounts of GZ17-8.20,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 38C is a graph of cell number versus dosage amounts of GZ17-8.20,illustrating the effect thereof in inducing the death of prostatecancer;

FIG. 38D is a graph of cell number versus dosage amounts of GZ17-8.20,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 38E is a graph of cell number versus dosage amounts of GZ17-8.20,illustrating the effect thereof in inducing the death of breast cancer;

FIG. 38F is a graph of cell number versus dosage amounts of GZ17-8.20,illustrating the effect thereof in inducing the death of leukemia;

FIG. 38G is a graph of cell number versus dosage amounts of GZ17-8.20,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 39A is a graph of cell number versus dosage amounts of GZ17-8.21,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 39B is a graph of cell number versus dosage amounts of GZ17-8.21,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 39C is a graph of cell number versus dosage amounts of GZ17-8.21,illustrating the effect thereof in inducing the death of prostatecancer;

FIG. 39D is a graph of cell number versus dosage amounts of GZ17-8.21,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 39E is a graph of cell number versus dosage amounts of GZ17-8.21,illustrating the effect thereof in inducing the death of breast cancer;

FIG. 39F is a graph of cell number versus dosage amounts of GZ17-8.21,illustrating the effect thereof in inducing the death of leukemia;

FIG. 39G is a graph of cell number versus dosage amounts of GZ17-8.21,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 40A is a graph of cell number versus dosage amounts of GZ17-8.22,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 40B is a graph of cell number versus dosage amounts of GZ17-8.22,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 40C is a graph of cell number versus dosage amounts of GZ17-8.22,illustrating the effect thereof in inducing the death of prostatecancer;

FIG. 40D is a graph of cell number versus dosage amounts of GZ17-8.22,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 40E is a graph of cell number versus dosage amounts of GZ17-8.22,illustrating the effect thereof in inducing the death of breast cancer;

FIG. 40F is a graph of cell number versus dosage amounts of GZ17-8.22,illustrating the effect thereof in inducing the death of leukemia;

FIG. 40G is a graph of cell number versus dosage amounts of GZ17-8.22,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 41A is a graph of cell number versus dosage amounts of GZ17-8.23,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 41B is a graph of cell number versus dosage amounts of GZ17-8.23,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 41C is a graph of cell number versus dosage amounts of GZ17-8.23,illustrating the effect thereof in inducing the death of prostatecancer;

FIG. 41D is a graph of cell number versus dosage amounts of GZ17-8.23,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 41E is a graph of cell number versus dosage amounts of GZ17-8.23,illustrating the effect thereof in inducing the death of breast cancer;

FIG. 41F is a graph of cell number versus dosage amounts of GZ17-8.23,illustrating the effect thereof in inducing the death of leukemia;

FIG. 41G is a graph of cell number versus dosage amounts of GZ17-8.23,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 42A is a graph of cell number versus dosage amounts of GZ17-8.24,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 42B is a graph of cell number versus dosage amounts of GZ17-8.24,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 42C is a graph of cell number versus dosage amounts of GZ17-8.24,illustrating the effect thereof in inducing the death of prostatecancer;

FIG. 42D is a graph of cell number versus dosage amounts of GZ17-8.24,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 42E is a graph of cell number versus dosage amounts of GZ17-8.24,illustrating the effect thereof in inducing the death of breast cancer;

FIG. 42F is a graph of cell number versus dosage amounts of GZ17-8.24,illustrating the effect thereof in inducing the death of leukemia;

FIG. 42G is a graph of cell number versus dosage amounts of GZ17-8.24,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 43A is a graph of cell number versus dosage amounts of GZ17-8.25,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 43B is a graph of cell number versus dosage amounts of GZ17-8.25,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 43C is a graph of cell number versus dosage amounts of GZ17-8.25,illustrating the effect thereof in inducing the death of prostatecancer;

FIG. 43D is a graph of cell number versus dosage amounts of GZ17-8.25,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 43E is a graph of cell number versus dosage amounts of GZ17-8.25,illustrating the effect thereof in inducing the death of breast cancer;

FIG. 43F is a graph of cell number versus dosage amounts of GZ17-8.25,illustrating the effect thereof in inducing the death of leukemia;

FIG. 43G is a graph of cell number versus dosage amounts of GZ17-8.25,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 44A is a graph of cell number versus dosage amounts of GZ17-8.26,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 44B is a graph of cell number versus dosage amounts of GZ17-8.26,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 44C is a graph of cell number versus dosage amounts of GZ17-8.26,illustrating the effect thereof in inducing the death of prostatecancer;

FIG. 44D is a graph of cell number versus dosage amounts of GZ17-8.26,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 44E is a graph of cell number versus dosage amounts of GZ17-8.26,illustrating the effect thereof in inducing the death of breast cancer;

FIG. 44F is a graph of cell number versus dosage amounts of GZ17-8.26,illustrating the effect thereof in inducing the death of leukemia;

FIG. 44G is a graph of cell number versus dosage amounts of GZ17-8.26,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 45A is a graph of cell number versus dosage amounts of GZ17-8.27,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 45B is a graph of cell number versus dosage amounts of GZ17-8.27,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 45C is a graph of cell number versus dosage amounts of GZ17-8.27,illustrating the effect thereof in inducing the death of prostatecancer;

FIG. 45D is a graph of cell number versus dosage amounts of GZ17-8.27,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 45E is a graph of cell number versus dosage amounts of GZ17-8.27,illustrating the effect thereof in inducing the death of breast cancer;

FIG. 45F is a graph of cell number versus dosage amounts of GZ17-8.27,illustrating the effect thereof in inducing the death of leukemia;

FIG. 45G is a graph of cell number versus dosage amounts of GZ17-8.27,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 46A is a graph of cell number versus dosage amounts of GZ17-8.28,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 46B is a graph of cell number versus dosage amounts of GZ17-8.28,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 46C is a graph of cell number versus dosage amounts of GZ17-8.28,illustrating the effect thereof in inducing the death of prostatecancer;

FIG. 46D is a graph of cell number versus dosage amounts of GZ17-8.28,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 46E is a graph of cell number versus dosage amounts of GZ17-8.28,illustrating the effect thereof in inducing the death of breast cancer;

FIG. 46F is a graph of cell number versus dosage amounts of GZ17-8.28,illustrating the effect thereof in inducing the death of leukemia;

FIG. 46G is a graph of cell number versus dosage amounts of GZ17-8.28,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 47A is a graph of cell number versus dosage amounts of GZ17-8.29,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 47B is a graph of cell number versus dosage amounts of GZ17-8.29,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 47C is a graph of cell number versus dosage amounts of GZ17-8.29,illustrating the effect thereof in inducing the death of prostatecancer;

FIG. 47D is a graph of cell number versus dosage amounts of GZ17-8.29,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 47E is a graph of cell number versus dosage amounts of GZ17-8.29,illustrating the effect thereof in inducing the death of breast cancer;

FIG. 47F is a graph of cell number versus dosage amounts of GZ17-8.29,illustrating the effect thereof in inducing the death of leukemia;

FIG. 47G is a graph of cell number versus dosage amounts of GZ17-8.29,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 48A is a graph of cell number versus dosage amounts of GZ17-8.30,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 48B is a graph of cell number versus dosage amounts of GZ17-8.30,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 48C is a graph of cell number versus dosage amounts of GZ17-8.30,illustrating the effect thereof in inducing the death of prostatecancer;

FIG. 48D is a graph of cell number versus dosage amounts of GZ17-8.30,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 48E is a graph of cell number versus dosage amounts of GZ17-8.30,illustrating the effect thereof in inducing the death of breast cancer;

FIG. 48F is a graph of cell number versus dosage amounts of GZ17-8.30,illustrating the effect thereof in inducing the death of leukemia;

FIG. 48G is a graph of cell number versus dosage amounts of GZ17-8.30,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 49A is a graph of cell number versus dosage amounts of GZ17-8.31,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 49B is a graph of cell number versus dosage amounts of GZ17-8.31,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 49C is a graph of cell number versus dosage amounts of GZ17-8.31,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 49D is a graph of cell number versus dosage amounts of GZ17-8.31,illustrating the effect thereof in inducing the death of leukemia;

FIG. 50A is a graph of cell number versus dosage amounts of GZ17-8.32,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 50B is a graph of cell number versus dosage amounts of GZ17-8.32,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 50C is a graph of cell number versus dosage amounts of GZ17-8.32,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 50D is a graph of cell number versus dosage amounts of GZ17-8.32,illustrating the effect thereof in inducing the death of leukemia;

FIG. 51A is a graph of cell number versus dosage amounts of GZ17-8.33,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 51B is a graph of cell number versus dosage amounts of GZ17-8.33,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 51C is a graph of cell number versus dosage amounts of GZ17-8.33,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 51D is a graph of cell number versus dosage amounts of GZ17-8.33,illustrating the effect thereof in inducing the death of leukemia;

FIG. 52A is a graph of cell number versus dosage amounts of GZ17-8.34,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 52B is a graph of cell number versus dosage amounts of GZ17-8.34,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 52C is a graph of cell number versus dosage amounts of GZ17-8.34,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 52D is a graph of cell number versus dosage amounts of GZ17-8.34,illustrating the effect thereof in inducing the death of leukemia;

FIG. 53A is a graph of cell number versus dosage amounts of GZ17-8.35,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 53B is a graph of cell number versus dosage amounts of GZ17-8.35,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 53C is a graph of cell number versus dosage amounts of GZ17-8.35,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 53D is a graph of cell number versus dosage amounts of GZ17-8.35,illustrating the effect thereof in inducing the death of leukemia;

FIG. 54A is a graph of cell number versus dosage amounts of GZ17-8.36,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 54B is a graph of cell number versus dosage amounts of GZ17-8.36,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 54C is a graph of cell number versus dosage amounts of GZ17-8.36,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 54D is a graph of cell number versus dosage amounts of GZ17-8.36,illustrating the effect thereof in inducing the death of leukemia;

FIG. 55A is a graph of cell number versus dosage amounts of GZ17-8.37,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 55B is a graph of cell number versus dosage amounts of GZ17-8.37,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 55C is a graph of cell number versus dosage amounts of GZ17-8.37,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 55D is a graph of cell number versus dosage amounts of GZ17-8.37,illustrating the effect thereof in inducing the death of leukemia;

FIG. 56A is a graph of cell number versus dosage amounts of GZ17-8.38,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 56B is a graph of cell number versus dosage amounts of GZ17-8.38,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 56C is a graph of cell number versus dosage amounts of GZ17-8.38,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 56D is a graph of cell number versus dosage amounts of GZ17-8.38,illustrating the effect thereof in inducing the death of leukemia;

FIG. 57A is a graph of cell number versus dosage amounts of GZ17-8.39,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 57B is a graph of cell number versus dosage amounts of GZ17-8.39,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 57C is a graph of cell number versus dosage amounts of GZ17-8.39,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 57D is a graph of cell number versus dosage amounts of GZ17-8.39,illustrating the effect thereof in inducing the death of leukemia;

FIG. 58A is a graph of cell number versus dosage amounts of GZ17-8.40,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 58B is a graph of cell number versus dosage amounts of GZ17-8.40,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 58C is a graph of cell number versus dosage amounts of GZ17-8.40,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 58D is a graph of cell number versus dosage amounts of GZ17-8.40,illustrating the effect thereof in inducing the death of leukemia;

FIG. 59A is a graph of cell number versus dosage amounts of GZ17-8.41,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 59B is a graph of cell number versus dosage amounts of GZ17-8.41,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 59C is a graph of cell number versus dosage amounts of GZ17-8.41,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 59D is a graph of cell number versus dosage amounts of GZ17-8.41,illustrating the effect thereof in inducing the death of leukemia;

FIG. 60A is a graph of cell number versus dosage amounts of GZ17-8.42,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 60B is a graph of cell number versus dosage amounts of GZ17-8.42,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 60C is a graph of cell number versus dosage amounts of GZ17-8.42,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 60D is a graph of cell number versus dosage amounts of GZ17-8.42,illustrating the effect thereof in inducing the death of leukemia;

FIG. 61A is a graph of cell number versus dosage amounts of GZ17-8.43,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 61B is a graph of cell number versus dosage amounts of GZ17-8.43,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 61C is a graph of cell number versus dosage amounts of GZ17-8.43,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 61D is a graph of cell number versus dosage amounts of GZ17-8.43,illustrating the effect thereof in inducing the death of leukemia;

FIG. 62A is a graph of cell number versus dosage amounts of GZ17-8.44,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 62B is a graph of cell number versus dosage amounts of GZ17-8.44,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 62C is a graph of cell number versus dosage amounts of GZ17-8.44,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 62D is a graph of cell number versus dosage amounts of GZ17-8.44,illustrating the effect thereof in inducing the death of leukemia;

FIG. 63A is a graph of cell number versus dosage amounts of GZ17-8.45,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 63B is a graph of cell number versus dosage amounts of GZ17-8.45,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 63C is a graph of cell number versus dosage amounts of GZ17-8.45,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 63D is a graph of cell number versus dosage amounts of GZ17-8.45,illustrating the effect thereof in inducing the death of leukemia;

FIG. 64A is a graph of cell number versus dosage amounts of GZ17-8.46,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 64B is a graph of cell number versus dosage amounts of GZ17-8.46,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 64C is a graph of cell number versus dosage amounts of GZ17-8.46,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 64D is a graph of cell number versus dosage amounts of GZ17-8.46,illustrating the effect thereof in inducing the death of leukemia;

FIG. 65A is a graph of cell number versus dosage amounts of GZ17-8.47,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 65B is a graph of cell number versus dosage amounts of GZ17-8.47,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 65C is a graph of cell number versus dosage amounts of GZ17-8.47,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 65D is a graph of cell number versus dosage amounts of GZ17-8.47,illustrating the effect thereof in inducing the death of leukemia;

FIG. 66A is a graph of cell number versus dosage amounts of GZ17-8.48,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 66B is a graph of cell number versus dosage amounts of GZ17-8.48,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 66C is a graph of cell number versus dosage amounts of GZ17-8.48,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 66D is a graph of cell number versus dosage amounts of GZ17-8.48,illustrating the effect thereof in inducing the death of leukemia;

FIG. 67A is a graph of cell number versus dosage amounts of GZ17-8.49,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 67B is a graph of cell number versus dosage amounts of GZ17-8.49,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 67C is a graph of cell number versus dosage amounts of GZ17-8.49,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 67D is a graph of cell number versus dosage amounts of GZ17-8.49,illustrating the effect thereof in inducing the death of leukemia;

FIG. 68A is a graph of cell number versus dosage amounts of GZ17-8.50,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 68B is a graph of cell number versus dosage amounts of GZ17-8.50,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 68C is a graph of cell number versus dosage amounts of GZ17-8.50,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 68D is a graph of cell number versus dosage amounts of GZ17-8.50,illustrating the effect thereof in inducing the death of leukemia;

FIG. 69A is a graph of cell number versus dosage amounts of GZ17-8.51,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 69B is a graph of cell number versus dosage amounts of GZ17-8.51,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 69C is a graph of cell number versus dosage amounts of GZ17-8.51,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 69D is a graph of cell number versus dosage amounts of GZ17-8.51,illustrating the effect thereof in inducing the death of leukemia;

FIG. 70A is a graph of cell number versus dosage amounts of GZ17-8.52,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 70B is a graph of cell number versus dosage amounts of GZ17-8.52,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 70C is a graph of cell number versus dosage amounts of GZ17-8.52,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 70D is a graph of cell number versus dosage amounts of GZ17-8.52,illustrating the effect thereof in inducing the death of leukemia;

FIG. 71A is a graph of cell number versus dosage amounts of GZ17-8.53,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 71B is a graph of cell number versus dosage amounts of GZ17-8.53,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 71C is a graph of cell number versus dosage amounts of GZ17-8.53,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 71D is a graph of cell number versus dosage amounts of GZ17-8.53,illustrating the effect thereof in inducing the death of leukemia;

FIG. 72A is a graph of cell number versus dosage amounts of GZ17-8.54,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 72B is a graph of cell number versus dosage amounts of GZ17-8.54,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 72C is a graph of cell number versus dosage amounts of GZ17-8.54,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 72D is a graph of cell number versus dosage amounts of GZ17-8.54,illustrating the effect thereof in inducing the death of leukemia;

FIG. 73A is a graph of cell number versus dosage amounts of GZ17-8.55,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 73B is a graph of cell number versus dosage amounts of GZ17-8.55,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 73C is a graph of cell number versus dosage amounts of GZ17-8.55,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 73D is a graph of cell number versus dosage amounts of GZ17-8.55,illustrating the effect thereof in inducing the death of leukemia;

FIG. 74A is a graph of cell number versus dosage amounts of GZ17-8.56,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 74B is a graph of cell number versus dosage amounts of GZ17-8.56,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 74C is a graph of cell number versus dosage amounts of GZ17-8.56,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 74D is a graph of cell number versus dosage amounts of GZ17-8.56,illustrating the effect thereof in inducing the death of leukemia;

FIG. 75A is a graph of cell number versus dosage amounts of GZ17-8.57,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 75B is a graph of cell number versus dosage amounts of GZ17-8.57,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 75C is a graph of cell number versus dosage amounts of GZ17-8.57,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 75D is a graph of cell number versus dosage amounts of GZ17-8.57,illustrating the effect thereof in inducing the death of leukemia;

FIG. 76A is a graph of cell number versus dosage amounts of GZ17-8.58,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 76B is a graph of cell number versus dosage amounts of GZ17-8.58,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 76C is a graph of cell number versus dosage amounts of GZ17-8.58,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 76D is a graph of cell number versus dosage amounts of GZ17-8.58,illustrating the effect thereof in inducing the death of leukemia;

FIG. 77A is a graph of cell number versus dosage amounts of GZ17-8.59,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 77B is a graph of cell number versus dosage amounts of GZ17-8.59,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 77C is a graph of cell number versus dosage amounts of GZ17-8.59,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 77D is a graph of cell number versus dosage amounts of GZ17-8.59,illustrating the effect thereof in inducing the death of leukemia;

FIG. 78A is a graph of cell number versus dosage amounts of GZ17-8.60,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 78B is a graph of cell number versus dosage amounts of GZ17-8.60,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 78C is a graph of cell number versus dosage amounts of GZ17-8.60,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 78D is a graph of cell number versus dosage amounts of GZ17-8.60,illustrating the effect thereof in inducing the death of leukemia;

FIG. 79A is a graph of cell number versus dosage amounts of GZ17-8.61,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 79B is a graph of cell number versus dosage amounts of GZ17-8.61,illustrating the effect thereof in inducing the death of lung cancer;

FIG. 79C is a graph of cell number versus dosage amounts of GZ17-8.61,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 79D is a graph of cell number versus dosage amounts of GZ17-8.61,illustrating the effect thereof in inducing the death of leukemia;

FIG. 80A is a graph of cell number versus dosage amounts of GZ17-10.04,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 80B is a graph of cell number versus dosage amounts of GZ17-10.05,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 80C is a graph of cell number versus dosage amounts of GZ17-10.06,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 80D is a graph of cell number versus dosage amounts of GZ17-10.04,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 80E is a graph of cell number versus dosage amounts of GZ17-10.05,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 80F is a graph of cell number versus dosage amounts of GZ17-10.06,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 80G is a comparative bar graph illustrating the comparativelymphoma cell-killing effect of the individual components,GZ17-10.04-10.06, versus the theoretical additive effect of thesecomponents, and the actual effect thereof, demonstrating the synergismof the three-component composition;

FIG. 80H is a comparative bar graph illustrating the comparativelymphoma cell-killing effect of the individual components, GZ17-10.04and 10.06, versus the theoretical additive effect of these components,and the actual effect thereof, demonstrating the synergism of thethree-component composition;

FIG. 80I is a graph of cell number versus dosage amounts of GZ17-10.04,illustrating the effect thereof in inducing the death of leukemia;

FIG. 80J is a graph of cell number versus dosage amounts of GZ17-10.05,illustrating the effect thereof in inducing the death of leukemia;

FIG. 80K is a graph of cell number versus dosage amounts of GZ17-10.06,illustrating the effect thereof in inducing the death of leukemia;

FIG. 80L is a comparative bar graph illustrating the comparativeleukemia cell-killing effect of the individual components,GZ17-10.04-10.06, versus the theoretical additive effect of thesecomponents, and the actual effect thereof, demonstrating the synergismof the three-component composition;

FIG. 80M is a comparative bar graph illustrating the comparativeleukemia cell-killing effect of the individual components, GZ17-10.04and 10.06, versus the theoretical additive effect of these components,and the actual effect thereof, demonstrating the synergism of thethree-component composition;

FIG. 80N is a graph of cell number versus dosage amounts of GZ17-10.04,illustrating the effect thereof in inducing the death of breast cancer;

FIG. 80O is a graph of cell number versus dosage amounts of GZ17-10.05,illustrating the effect thereof in inducing the death of breast cancer;

FIG. 80P is a graph of cell number versus dosage amounts of GZ17-10.06,illustrating the effect thereof in inducing the death of breast cancer;

FIG. 80Q is a comparative bar graph illustrating the comparative breastcancer cell-killing effect of the individual components,GZ17-10.04-10.06, versus the theoretical additive effect of thesecomponents, and the actual effect thereof, demonstrating the synergismof the three-component composition;

FIG. 80R is a comparative bar graph illustrating the comparative breastcancer cell-killing effect of the individual components, GZ17-10.04 and10.06, versus the theoretical additive effect of these components, andthe actual effect thereof, demonstrating the synergism of thethree-component composition;

FIG. 81A is a graph of cell number versus dosage amounts of GZ17-08.512,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 81B is a graph of cell number versus dosage amounts of GZ17-08.512,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 81C is a graph of cell number versus dosage amounts of GZ17-08.512,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 81D is a graph of cell number versus dosage amounts of GZ17-08.513,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 81E is a graph of cell number versus dosage amounts of GZ17-08.513,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 81F is a graph of cell number versus dosage amounts of GZ17-08.513,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 81G is a graph of cell number versus dosage amounts of GZ17-08.514,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 81H is a graph of cell number versus dosage amounts of GZ17-08.514,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 81I is a graph of cell number versus dosage amounts of GZ17-08.514,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 81J is a graph of cell number versus dosage amounts of GZ17-08.515,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 81K is a graph of cell number versus dosage amounts of GZ17-08.515,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 81L is a graph of cell number versus dosage amounts of GZ17-08.515,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 81M is a graph of cell number versus dosage amounts of GZ17-08.516,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 81N is a graph of cell number versus dosage amounts of GZ17-08.516,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 81O is a graph of cell number versus dosage amounts of GZ17-08.516,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 81P is a graph of cell number versus dosage amounts of GZ17-08.517,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 81Q is a graph of cell number versus dosage amounts of GZ17-08.517,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 81R is a graph of cell number versus dosage amounts of GZ17-08.517,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 81S is a graph of cell number versus dosage amounts of GZ17-08.518,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 81T is a graph of cell number versus dosage amounts of GZ17-08.518,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 81U is a graph of cell number versus dosage amounts of GZ17-08.518,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 81V is a graph of cell number versus dosage amounts of GZ17-08.519,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 81W is a graph of cell number versus dosage amounts of GZ17-08.519,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 81X is a graph of cell number versus dosage amounts of GZ17-08.519,illustrating the effect thereof in inducing the death of head and neckcancer;

FIG. 81Y is a graph of cell number versus dosage amounts of GZ17-08.520,illustrating the effect thereof in inducing the death of ovarian cancer;

FIG. 81Z is a graph of cell number versus dosage amounts of GZ17-08.520,illustrating the effect thereof in inducing the death of lymphoma;

FIG. 81AA is a graph of cell number versus dosage amounts ofGZ17-08.520, illustrating the effect thereof in inducing the death ofhead and neck cancer;

FIG. 81BB is a graph of cell number versus dosage amounts ofGZ17-08.521, illustrating the effect thereof in inducing the death ofovarian cancer;

FIG. 81CC is a graph of cell number versus dosage amounts ofGZ17-08.521, illustrating the effect thereof in inducing the death oflymphoma;

FIG. 81DD is a graph of cell number versus dosage amounts ofGZ17-08.521, illustrating the effect thereof in inducing the death ofhead and neck cancer;

FIG. 82-1 is a graph of cell number versus dosage amounts of GZ08.065,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-2 is a graph of cell number versus dosage amounts of GZ08.066,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-3 is a graph of cell number versus dosage amounts of GZ08.067,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-4 is a graph of cell number versus dosage amounts of GZ08.068,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-5 is a graph of cell number versus dosage amounts of GZ08.069,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-6 is a graph of cell number versus dosage amounts of GZ08.070,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-7 is a graph of cell number versus dosage amounts of GZ08.071,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-8 is a graph of cell number versus dosage amounts of GZ08.072,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-9 is a graph of cell number versus dosage amounts of GZ08.073,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-10 is a graph of cell number versus dosage amounts of GZ08.074,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-11 is a graph of cell number versus dosage amounts of GZ08.075,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-12 is a graph of cell number versus dosage amounts of GZ08.076,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-13 is a graph of cell number versus dosage amounts of GZ08.077,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-14 is a graph of cell number versus dosage amounts of GZ08.078,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-15 is a graph of cell number versus dosage amounts of GZ08.079,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-16 is a graph of cell number versus dosage amounts of GZ08.080,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-17 is a graph of cell number versus dosage amounts of GZ08.081,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-18 is a graph of cell number versus dosage amounts of GZ08.082,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-19 is a graph of cell number versus dosage amounts of GZ08.083,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-20 is a graph of cell number versus dosage amounts of GZ08.084,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-21 is a graph of cell number versus dosage amounts of GZ08.085,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-22 is a graph of cell number versus dosage amounts of GZ08.086,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-23 is a graph of cell number versus dosage amounts of GZ08.087,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-24 is a graph of cell number versus dosage amounts of GZ08.088,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-25 is a graph of cell number versus dosage amounts of GZ08.089,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-26 is a graph of cell number versus dosage amounts of GZ08.090,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-27 is a graph of cell number versus dosage amounts of GZ08.091,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-28 is a graph of cell number versus dosage amounts of GZ08.092,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-29 is a graph of cell number versus dosage amounts of GZ08.093,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-30 is a graph of cell number versus dosage amounts of GZ08.094,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-31 is a graph of cell number versus dosage amounts of GZ08.095,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-32 is a graph of cell number versus dosage amounts of GZ08.096,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-33 is a graph of cell number versus dosage amounts of GZ08.097,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-34 is a graph of cell number versus dosage amounts of GZ08.098,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-35 is a graph of cell number versus dosage amounts of GZ08.099,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-36 is a graph of cell number versus dosage amounts of GZ08.100,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-37 is a graph of cell number versus dosage amounts of GZ08.101,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-38 is a graph of cell number versus dosage amounts of GZ08.102,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-39 is a graph of cell number versus dosage amounts of GZ08.103,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-40 is a graph of cell number versus dosage amounts of GZ08.104,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-41 is a graph of cell number versus dosage amounts of GZ08.105,illustrating the effect thereof in inducing the death of lung cancer, asdescribed in Example 82;

FIG. 82-42 is a graph of cell number versus dosage amounts of GZ08.065,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-43 is a graph of cell number versus dosage amounts of GZ08.066,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-44 is a graph of cell number versus dosage amounts of GZ08.067,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-45 is a graph of cell number versus dosage amounts of GZ08.068,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-46 is a graph of cell number versus dosage amounts of GZ08.069,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-47 is a graph of cell number versus dosage amounts of GZ08.070,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-48 is a graph of cell number versus dosage amounts of GZ08.071,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-49 is a graph of cell number versus dosage amounts of GZ08.072,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-50 is a graph of cell number versus dosage amounts of GZ08.073,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-51 is a graph of cell number versus dosage amounts of GZ08.074,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-52 is a graph of cell number versus dosage amounts of GZ08.075,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-53 is a graph of cell number versus dosage amounts of GZ08.076,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-54 is a graph of cell number versus dosage amounts of GZ08.077,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-55 is a graph of cell number versus dosage amounts of GZ08.078,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-56 is a graph of cell number versus dosage amounts of GZ08.079,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-57 is a graph of cell number versus dosage amounts of GZ08.080,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-58 is a graph of cell number versus dosage amounts of GZ08.081,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-59 is a graph of cell number versus dosage amounts of GZ08.082,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-60 is a graph of cell number versus dosage amounts of GZ08.083,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-61 is a graph of cell number versus dosage amounts of GZ08.084,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-62 is a graph of cell number versus dosage amounts of GZ08.085,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-63 is a graph of cell number versus dosage amounts of GZ08.086,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-64 is a graph of cell number versus dosage amounts of GZ08.087,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-65 is a graph of cell number versus dosage amounts of GZ08.088,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-66 is a graph of cell number versus dosage amounts of GZ08.089,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-67 is a graph of cell number versus dosage amounts of GZ08.090,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-68 is a graph of cell number versus dosage amounts of GZ08.091,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-69 is a graph of cell number versus dosage amounts of GZ08.092,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-70 is a graph of cell number versus dosage amounts of GZ08.093,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-71 is a graph of cell number versus dosage amounts of GZ08.094,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-72 is a graph of cell number versus dosage amounts of GZ08.095,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-73 is a graph of cell number versus dosage amounts of GZ08.096,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-74 is a graph of cell number versus dosage amounts of GZ08.097,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-75 is a graph of cell number versus dosage amounts of GZ08.098,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-76 is a graph of cell number versus dosage amounts of GZ08.099,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-77 is a graph of cell number versus dosage amounts of GZ08.100,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-78 is a graph of cell number versus dosage amounts of GZ08.101,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-79 is a graph of cell number versus dosage amounts of GZ08.102,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-80 is a graph of cell number versus dosage amounts of GZ08.103,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-81 is a graph of cell number versus dosage amounts of GZ08.104,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-82 is a graph of cell number versus dosage amounts of GZ08.105,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 82;

FIG. 82-83 is a graph of cell number versus dosage amounts of GZ08.065,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-84 is a graph of cell number versus dosage amounts of GZ08.066,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-85 is a graph of cell number versus dosage amounts of GZ08.067,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-86 is a graph of cell number versus dosage amounts of GZ08.068,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-87 is a graph of cell number versus dosage amounts of GZ08.069,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-88 is a graph of cell number versus dosage amounts of GZ08.070,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-89 is a graph of cell number versus dosage amounts of GZ08.071,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-90 is a graph of cell number versus dosage amounts of GZ08.072,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-91 is a graph of cell number versus dosage amounts of GZ08.073,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-92 is a graph of cell number versus dosage amounts of GZ08.074,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-93 is a graph of cell number versus dosage amounts of GZ08.075,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-94 is a graph of cell number versus dosage amounts of GZ08.076,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-95 is a graph of cell number versus dosage amounts of GZ08.077,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-96 is a graph of cell number versus dosage amounts of GZ08.078,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-97 is a graph of cell number versus dosage amounts of GZ08.079,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-98 is a graph of cell number versus dosage amounts of GZ08.080,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-99 is a graph of cell number versus dosage amounts of GZ08.081,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-100 is a graph of cell number versus dosage amounts of GZ08.082,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-101 is a graph of cell number versus dosage amounts of GZ08.083,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-102 is a graph of cell number versus dosage amounts of GZ08.084,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-103 is a graph of cell number versus dosage amounts of GZ08.085,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-104 is a graph of cell number versus dosage amounts of GZ08.086,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-105 is a graph of cell number versus dosage amounts of GZ08.087,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-106 is a graph of cell number versus dosage amounts of GZ08.088,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-107 is a graph of cell number versus dosage amounts of GZ08.089,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-108 is a graph of cell number versus dosage amounts of GZ08.090,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-109 is a graph of cell number versus dosage amounts of GZ08.091,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-110 is a graph of cell number versus dosage amounts of GZ08.092,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-111 is a graph of cell number versus dosage amounts of GZ08.093,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-112 is a graph of cell number versus dosage amounts of GZ08.094,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-113 is a graph of cell number versus dosage amounts of GZ08.095,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-114 is a graph of cell number versus dosage amounts of GZ08.096,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-115 is a graph of cell number versus dosage amounts of GZ08.097,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-116 is a graph of cell number versus dosage amounts of GZ08.098,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-117 is a graph of cell number versus dosage amounts of GZ08.099,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-118 is a graph of cell number versus dosage amounts of GZ08.100,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-119 is a graph of cell number versus dosage amounts of GZ08.101,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-120 is a graph of cell number versus dosage amounts of GZ08.102,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-121 is a graph of cell number versus dosage amounts of GZ08.103,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-122 is a graph of cell number versus dosage amounts of GZ08.104,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-123 is a graph of cell number versus dosage amounts of GZ08.105,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 82;

FIG. 82-124 is a comparative bar graph illustrating the comparative lungcancer cell-killing effect of the individual components of GZ08.065,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-125 is a comparative bar graph illustrating the comparative lungcancer cell-killing effect of the individual components of GZ08.067,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-126 is a comparative bar graph illustrating the comparative lungcancer cell-killing effect of the individual components of GZ08.068,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-127 is a comparative bar graph illustrating the comparative lungcancer cell-killing effect of the individual components of GZ08.079,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-128 is a comparative bar graph illustrating the comparative lungcancer cell-killing effect of the individual components of GZ08.080,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition;

FIG. 82-129 is a comparative bar graph illustrating the comparative lungcancer cell-killing effect of the individual components of GZ08.084,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-130 is a comparative bar graph illustrating the comparative lungcancer cell-killing effect of the individual components of GZ08.085,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-131 is a comparative bar graph illustrating the comparative lungcancer cell-killing effect of the individual components of GZ08.086,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-132 is a comparative bar graph illustrating the comparative lungcancer cell-killing effect of the individual components of GZ08.087,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-133 is a comparative bar graph illustrating the comparative lungcancer cell-killing effect of the individual components of GZ08.088,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-134 is a comparative bar graph illustrating the comparative lungcancer cell-killing effect of the individual components of GZ08.089,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-135 is a comparative bar graph illustrating the comparative lungcancer cell-killing effect of the individual components of GZ08.090,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-136 is a comparative bar graph illustrating the comparative lungcancer cell-killing effect of the individual components of GZ08.091,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-137 is a comparative bar graph illustrating the comparative lungcancer cell-killing effect of the individual components of GZ08.092,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-138 is a comparative bar graph illustrating the comparative lungcancer cell-killing effect of the individual components of GZ08.093,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-139 is a comparative bar graph illustrating the comparative lungcancer cell-killing effect of the individual components of GZ08.094,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-140 is a comparative bar graph illustrating the comparative lungcancer cell-killing effect of the individual components of GZ08.097,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-141 is a comparative bar graph illustrating the comparative lungcancer cell-killing effect of the individual components of GZ08.098,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-142 is a comparative bar graph illustrating the comparative lungcancer cell-killing effect of the individual components of GZ08.100,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-143 is a comparative bar graph illustrating the comparative lungcancer cell-killing effect of the individual components of GZ08.101,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-144 is a comparative bar graph illustrating the comparative lungcancer cell-killing effect of the individual components of GZ08.102,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-145 is a comparative bar graph illustrating the comparative lungcancer cell-killing effect of the individual components of GZ08.103,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-146 is a comparative bar graph illustrating the comparative lungcancer cell-killing effect of the individual components of GZ08.104,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-147 is a comparative bar graph illustrating the comparative lungcancer cell-killing effect of the individual components of GZ08.105,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-148 is a comparative bar graph illustrating the comparativelymphoma cell-killing effect of the individual components of GZ08.067,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-149 is a comparative bar graph illustrating the comparativelymphoma cell-killing effect of the individual components of GZ08.073,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-150 is a comparative bar graph illustrating the comparativelymphoma cell-killing effect of the individual components of GZ08.076,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-151 is a comparative bar graph illustrating the comparativelymphoma cell-killing effect of the individual components of GZ08.077,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-152 is a comparative bar graph illustrating the comparativelymphoma cell-killing effect of the individual components of GZ08.080,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-153 is a comparative bar graph illustrating the comparativelymphoma cell-killing effect of the individual components of GZ08.083,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-154 is a comparative bar graph illustrating the comparativelymphoma cell-killing effect of the individual components of GZ08.086,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-155 is a comparative bar graph illustrating the comparativelymphoma cell-killing effect of the individual components of GZ08.088,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-156 is a comparative bar graph illustrating the comparativelymphoma cell-killing effect of the individual components of GZ08.092,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-157 is a comparative bar graph illustrating the comparativelymphoma cell-killing effect of the individual components of GZ08.095,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-158 is a comparative bar graph illustrating the comparativelymphoma cell-killing effect of the individual components of GZ08.099,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-159 is a comparative bar graph illustrating the comparativelymphoma cell-killing effect of the individual components of GZ08.101,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-160 is a comparative bar graph illustrating the comparativelymphoma cell-killing effect of the individual components of GZ08.105,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-161 is a comparative bar graph illustrating the comparativeleukemia cell-killing effect of the individual components of GZ08.086,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-162 is a comparative bar graph illustrating the comparativeleukemia cell-killing effect of the individual components of GZ08.087,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-163 is a comparative bar graph illustrating the comparativeleukemia cell-killing effect of the individual components of GZ08.090,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-164 is a comparative bar graph illustrating the comparativeleukemia cell-killing effect of the individual components of GZ08.094,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-165 is a comparative bar graph illustrating the comparativeleukemia cell-killing effect of the individual components of GZ08.098,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-166 is a comparative bar graph illustrating the comparativeleukemia cell-killing effect of the individual components of GZ08.104,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 82-167 is a comparative bar graph illustrating the comparativeleukemia cell-killing effect of the individual components of GZ08.105,versus the theoretical additive effect of these components, and theactual effect thereof at the indicted concentration, demonstrating thesynergism of the three-component composition, as described in Example82;

FIG. 83A is a graph of cell number versus dosage amounts oforthovanillin, illustrating the effect thereof in inducing the death oflymphoma, as described in Example 83;

FIG. 83B is a graph of cell number versus dosage amounts of curcumin,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 83;

FIG. 83C is a graph of cell number versus dosage amounts of harmaline,illustrating the effect thereof in inducing the death of lymphoma, asdescribed in Example 83;

FIG. 83D is a comparative bar graph illustrating the comparativelymphoma cell-killing effect of the individual components,orthovanillin, curcumin, and harmaline, as shown in FIGS. 83A-83C,versus the theoretical additive effect of these components, and theactual effect thereof, demonstrating the synergism of thethree-component composition;

FIG. 83E is a graph of cell number versus dosage amounts oforthovanillin, illustrating the effect thereof in inducing the death ofleukemia, as described in Example 83;

FIG. 83F is a graph of cell number versus dosage amounts of curcumin,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 83;

FIG. 83G is a graph of cell number versus dosage amounts of harmaline,illustrating the effect thereof in inducing the death of leukemia, asdescribed in Example 83;

FIG. 83H is a comparative bar graph illustrating the comparativeleukemia cell-killing effect of the individual components,orthovanillin, curcumin, and harmaline, as shown in FIGS. 83E-83G,versus the theoretical additive effect of these components, and theactual effect thereof, demonstrating the synergism of thethree-component composition;

FIG. 83I is a graph of cell number versus dosage amounts oforthovanillin, illustrating the effect thereof in inducing the death ofbreast cancer, as described in Example 83;

FIG. 83J is a graph of cell number versus dosage amounts of curcumin,illustrating the effect thereof in inducing the death of breast cancer,as described in Example 83;

FIG. 83K is a graph of cell number versus dosage amounts of harmaline,illustrating the effect thereof in inducing the death of breast cancer,as described in Example 83;

FIG. 83L is a comparative bar graph illustrating the comparative breastcancer cell-killing effect of the individual components, orthovanillin,curcumin, and harmaline, as shown in FIGS. 83I-83K, versus thetheoretical additive effect of these components, and the actual effectthereof, demonstrating the synergism of the three-component composition;

FIG. 84A is series of photographs of pancreatic cancer spheres includinga control, treatment of the spheres with GZ17-6.02 at levels of 0.5IC₅₀and IC₅₀, illustrating the effect of GZ17-6.02 in reducing the numberand size of pancreatic cancer spheres, as set forth in Example 84;

FIG. 84B is a series of blots illustrating in vitro pancreatic celltreatment with GZ17-6.02 at levels of 0.5IC₅₀ and IC₅₀, and depictingdecreases in cancer stem cell markers owing to the treatment withGZ17-6.02, as set forth in Example 84;

FIG. 84C is a series of blots illustrating the treatment of mousepancreatic cancer cells with GZ17-6.02, and depicting the decrease incancer stem cell markers owing to the treatment with GZ17-6.02, as setforth in Example 84; and

FIG. 84D is a possible pathway diagram illustrating the action ofGZ17-6.02 on both cancer stem cells (CSC) and other cancer cells withina tumor, as set forth in Example 84.

DETAILED DESCRIPTION

The therapeutic agents of the invention are used in therapeuticallyeffective amounts, i.e., amounts that will elicit the biological ormedical response of a tissue, system, or subject that is being sought,and in particular to elicit some desired therapeutic effect against avariety of human diseases, and especially cancers; in the case ofcancers, the agents operate by preventing and/or inhibitingproliferation and/or survival of cancerous cells, and/or by slowing theprogression of cancers. Those skilled in the art recognize that anamount may be considered therapeutically effective even if the conditionis not totally eradicated or prevented, but it or its symptoms and/oreffects are improved or alleviated partially in the subject. Of course,the appropriate makeup of the agents hereof and dosing regimens usingsuch agents will depend on the particular cancer being treated, theextent of the disease, and other factors related to the patient asdetermined by those skilled in the art. Hence, the terms “therapeutic”or “treat,” as used herein, refer to products or processes in accordancewith the invention that are intended to produce a beneficial change inan existing condition (e.g., cancerous tissue, tumor size, metastases,etc.) of a subject, such as by reducing the severity of the clinicalsymptoms and/or effects of the condition, and/or reducing the durationof the symptoms/effects of a subject.

Additional ingredients may be included with the chemotherapeutic agentsof the invention for administration to the subject. Such additionalingredients include, other active agents, preservatives, bufferingagents, salts, carriers, excipients, diluents, or otherpharmaceutically-acceptable ingredients. The active agents that could beincluded in the compositions include antiviral, antibiotic, or otheranticancer compounds.

The combined therapeutic agents of the invention preferably givesynergistic results, which are entirely unexpected. Moreover, the lackof side effects when the agents are administered to patients is quitesurprising and essentially unique. As used herein, the terms“combination” or “in combination” are intended to embrace compositionswherein the components are physically intermixed as dosage forms, and tosituations where the individual components are separately administeredto a subject over relatively short periods of time, which would have thesame therapeutic effects as a single dosage form.

In use, a therapeutically effective amount of an agent in accordancewith the invention is administered to a subject in need thereof. Suchmay comprise a single unit dosage or, more usually, periodic (e.g.,daily) administration of lower dosages over time. Advantageously,administration of such therapeutically effective amounts achieves anunexpected therapeutic synergy. This means that the therapeutic two- orthree-component compositions of the invention exhibit a joint actionwhere one or more of the components supplements or enhances the actionof at least one of the other components to produce an effect greaterthan that which may be obtained by use of individual components inequivalent quantities, or produce effects that could not be obtainedwith safe quantities of the other components, individually or incombination. Generally, one or more of the components working togetherproduce an effect greater than the sum of their individual effects.

The dosages may be administered in any convenient manner, such as byoral, rectal, nasal, ophthalmic, parenteral (including intraperitoneal,gastrointestinal, intrathecal, intravenous, cutaneous (e.g., dermalpatch), subcutaneous (e.g. injection or implant), or intramuscular)administrations. The dosage forms of the invention may be in the form ofliquids, gels, suspensions, solutions, or solids (e.g., tablets, pills,or capsules). Moreover, therapeutically effective amounts of the agentsof the invention may be co-administered with other chemotherapeuticagent(s), where the two products are administered substantiallysimultaneously or in any sequential manner.

Additional advantages of the various embodiments of the invention willbe apparent to those skilled in the art upon review of the disclosureherein and the working examples below. It will be appreciated that thevarious embodiments described herein are not necessarily mutuallyexclusive unless otherwise indicated herein. For example, a featuredescribed or depicted in one embodiment may also be included in otherembodiments, but is not necessarily included. Thus, the presentinvention encompasses a variety of combinations and/or integrations ofthe specific embodiments described herein.

As used herein, the phrase “and/or,” when used in a list of two or moreitems, means that any one of the listed items can be employed by itselfor any combination of two or more of the listed items can be employed.For example, if a composition is described as containing or excludingcomponents A, B, and/or C, the composition can contain or exclude Aalone; B alone; C alone; A and B in combination; A and C in combination;B and C in combination; or A, B, and C in combination.

The present description also uses numerical ranges to quantify certainparameters relating to various embodiments of the invention. It shouldbe understood that when numerical ranges are provided, such ranges areto be construed as providing literal support for claim limitations thatonly recite the lower value of the range as well as claim limitationsthat only recite the upper value of the range. For example, a disclosednumerical range of about 10 to about 100 provides literal support for aclaim reciting “greater than about 10” (with no upper bounds) and aclaim reciting “less than about 100” (with no lower bounds).

In the ensuing discussion, the curcumin, harmine, and isovanillincomponent(s) will be individually described. In such discussions, whereterms are used which specify or imply the presence of carbon-carbonchains (e.g., alkyl, alkenyl, alkoxy, alkyl amine, alkenyl amine,aldehyde, carboxylate, or the like), these disclosures should beunderstood to refer to primary (straight), branched chain, or cycliccarbon chain groups. Moreover, unless indicated otherwise, reference toaryl groups means phenyl, substituted phenyl, naphthyl, substitutednaphthyl; and heteroatom aryl groups refers to aryl groups containing anitrogen, oxygen, boron, or sulfur atom, such as pyridine; heterocyclicgroups refers to cyclic groups containing from 3-7 atoms, one or more ofwhich is a nitrogen, oxygen, boron, or sulfur heteroatom; and aminerefers to primary, secondary, tertiary, or quaternary amines.

As used herein, pharmaceutically acceptable salts with reference to thecomponents means salts of the component compounds of the presentinvention which are pharmaceutically acceptable, i.e., salts which areuseful in preparing pharmaceutical compositions that are generally safe,non-toxic, and neither biologically nor otherwise undesirable and areacceptable for human pharmaceutical use, and which possess the desireddegree of pharmacological activity. Such pharmaceutically acceptablesalts include acid addition salts formed with inorganic acids such ashydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid,phosphoric acid, and the like; or with organic acids such as1,2-ethanedisulfonic acid, 2-hydroxyethanesulfonic acid,2-naphthalenesulfonic acid, 3-phenylpropionic acid,4,4′-methylenebis(3-hydroxy-2-ene-1-carboxylic acid),4-methylbicyclo[2.2.2]oct-2-ene-1-carboxylic acid, acetic acid,aliphatic mono- and dicarboxylic acids, aliphatic sulfuric acids,aromatic sulfuric acids, benzenesulfonic acid, benzoic acid,camphorsulfonic acid, carbonic acid, cinnamic acid, citric acid,cyclopentanepropionic acid, ethanesulfonic acid, fumaric acid,glucoheptonic acid, gluconic acid, glutamic acid, glycolic acid,heptanoic acid, hexanoic acid, hydroxynaphthoic acid, lactic acid,laurylsulfuric acid, maleic acid, malic acid, malonic acid, mandelicacid, methanesulfonic acid, muconic acid, o-(4-hydroxybenzoyl)benzoicacid, oxalic acid, p-chlorobenzenesulfonic acid, phenyl-substitutedalkanoic acids, propionic acid, p-toluenesulfonic acid, pyruvic acid,salicylic acid, stearic acid, succinic acid, tartaric acid,tertiarybutylacetic acid, trimethylacetic acid, and the like.Pharmaceutically acceptable salts also include base addition salts whichmay be formed when acidic protons present are capable of reacting withinorganic or organic bases. Acceptable inorganic bases include sodiumhydroxide, sodium carbonate, potassium hydroxide, aluminum hydroxide andcalcium hydroxide. Acceptable organic bases include ethanolamine,diethanolamine, triethanolamine, tromethamine, N-methylglucamine and thelike. It should be recognized that the particular anion or cationforming a part of any salt of this invention is not critical, so long asthe salt, as a whole, is pharmacologically acceptable. Additionalexamples of pharmaceutically acceptable salts and their methods ofpreparation and use are presented in Handbook of Pharmaceutical SaltsProperties, and Use, P. H. Stahl & C. G. Wermuth eds., ISBN978-3-90639-058-1 (2008).

As noted below, the curcumin, harmine, and isovanillin component(s) maybe obtained as synthetic compounds of high purity, or from modifiednaturally occurring sources. In either case, however, it is preferredthat the component(s) be purified to a level of at least about 50% byweight, more preferably at least about 70% by weight, still morepreferably at least about 90% by weight, and most preferably at leastabout 98% by weight.

The individual discussions of the component(s) contain structuralformulas. In order to be entirely clear, curcumin-based formulas areindicated “C-number,” (not to be confused with carbon chain numbers,which are indicated as “Cnumber,” without an intervening hyphen),harmine-based formulas are indicated as “H-number,” andisovanillin-based formulas are indicated as “I-number.”

Before discussing the individual components of the invention, it shouldbe understood that use of unmodified, naturally occurring sources of thecomponents is generally not appropriate or desirable, because thesenaturally occurring products contain relatively small amounts of thedesired components and/or have potentially interfering compoundstherein. For example, naturally occurring turmeric has onlyapproximately 2-3% by weight curcumin therein and accordingly thestraightforward use of unmodified turmeric would not be suitable for theinvention. In like manner, naturally occurring harmala seed containsonly a very minor amount of harmine and such a product would also beinappropriate.

Thus, the preferred components of the invention are either syntheticallyderived or derived from one or more naturally occurring product(s) whichhave been significantly modified so as to contain at least about 25% byweight (more preferably at least about 50% by weight, and still morepreferably about 70% by weight) of the desired component. As usedherein, “synthetically derived” means that the component in question wassynthesized using specific starting ingredients and one or more chemicaland/or biological reactions to obtain substantially pure compounds.Modification of naturally occurring products may involve extractions, orany other physical or chemical steps to achieve the desired end product,e.g., harmine components may be obtained from treatment of harmala seed,or curcumin components may be obtained from treatment of turmeric.

For example, curcumin can be synthetically derived to a high degree ofpurity. Alternately, curcumin can be obtained by extraction or othertreatment of naturally occurring turmeric so that the curcumin contentof the modified turmeric has the above-noted levels of curcumin therein.

The Curcumin Component(s)

As used herein, “curcumin component(s)” shall mean curcumin, itsmetabolites and derivatives, isomers and tautomers thereof, esters,metal complexes (e.g., Cu, Fe, Zn, Pt, V), and pharmaceuticallyacceptable salts of any of the foregoing. Curcumin derivatives includeboth naturally occurring and synthetic derivatives, e.g., thespontaneous degradation products of curcumin, curcumin metabolites, andsynthetic curcumin derivative compounds.

1. Curcumin.

Curcumin (diferuloylmethane,1,7-bis(4-hydroxy3-mcthoxyphenyl)-1,6-heptadiene-3,5-dione) is asymmetrical diphenolic dienone, see structure C-1 below. It exists insolution as an equilibrium mixture of the symmetrical dienone (diketo)and the keto-enol tautomer; the keto-enol form is strongly favored byintramolecular hydrogen bonding.

Curcumin contains two aryl rings separated by an unsaturated 7-carbonlinker having a symmetrical β-diketone group (as used herein,“β-diketone” embraces both tautomeric forms, namely the diketo and enolforms). The aryl rings of curcumin contain a hydroxyl group in the paraposition and a methoxy group in the meta position.

2. Degradation Products of Curcumin.

It is known that, under certain pH and other conditions, curcumin willspontaneously form degradation products, and especially one or more ofthe following:

3. Curcumin Metabolites

It has been determined that curcumin is differently metabolized in vivodepending upon the route of administration, see, Shen et al. ThePharmacology of Curcumin: Is it the Degradation Products? Trends inMolecular Medicine, March 2012 Vol. 18, No. 2, incorporated by referenceherein it its entirety. Thus, when curcumin is orally administered, themetabolites normally include one or more of the following:

On the other hand, where the route of administration isintravenous/intraperitoneal, the metabolites generally include thefollowing:

C-9

Other naturally occurring curcumin derivatives include cyclocurcumin,bisdemethoxycurcumin, demethoxycurcumin, dihydrocurcumin, caffeic acid,cinnamic acid, isoeugenol, dibenzoylmethane, dehydrozingerone,capsaicin, [6]-gingerol, [6]-paradol, chlorogenic acid, yakuchinone A,oregonin, cassumuin A, and cassumuin B.

4. Synthetic Curcumin Derivatives.

Curcumin derivatives are expected to be beneficial for use in thetreatment methods of the invention. The term “curcumin derivative” isused interchangeably with the term “curcumin analog” and “curcuminanalogue” (alternative spelling) and includes, for example, curcuminderivatives, analogs, curcuminoids and chalcones. In one embodiment thecurcumin derivative includes first and second aryl groups covalentlyattached by way of a linker or a linking group. In another embodiment,the second aryl group is absent, such that the curcumin derivativecontains a first aryl group and the linker but no second aryl group atthe distal end of the linker. Optionally, the first and/or second arylgroup is a heteroaryl group. The first and second aryl groups may beindependently substituted or unsubstituted.

Curcumin derivatives that exhibit improved pharmacokinetic propertiesand/or reduced toxicity are preferred. For example, curcumin derivativesthat include heteroaryl groups and/or unsaturated linkers arc expectedto impart improved pharmacokinetic properties and/or reduced toxicity tothe compounds, because they are expected to be less chemically reactivein vivo. One example of preferred curcumin derivatives includes thoseincluding one or two carbonyl groups in the linker region, includingthose derivatives that preserve the enone functionality of curcumin.Derivatives that include heteroaryl groups and/or unsaturated linkersare expected to be less likely to be degraded and/or form toxic adductsor intermediates under physiological conditions.

Thus, in one aspect, curcumin derivatives of the invention are generallyencompassed by the formula:Ar1-L-Ar2  C-12where Ar1 and Ar2 are independently aryl groups, and L is a divalentlinking group that includes between 3 and 7 backbone carbon atoms,wherein one or more of the backbone carbon atoms include a carbonyl orhydroxyl moiety.

a. Aryl Groups

Preferred aryl groups include phenyl, naphthyl, thienyl, pyridinium, andpyridyl groups.

Aryl groups Ar1 and Ar2 may be substituted or unsubstituted, and one ormore of the ring carbons may be substituted with a heteroatom, andespecially N, S, B, or O.

For example, in one embodiment of the invention, Ar1 can be an arylgroup according to the formula:

and Ar2 can be an aryl group according to the formula:

where one or more of the aryl ring carbons of Ar1 and Ar2 may beindependently substituted with a heteroatom selected from N, S, B, or O,and R1-R10 are independently selected from the group consisting of H,hydroxyl, halogen, amine, nitro, sulfonate, sulfoxide, thio, ester,carboxylate, amide, borate, C1-C4 boronate, C1-C8 alkyl, C2-C8 alkenyl,C1-C6 haloalkyl, C1-C6 alkoxy, C1-C6 amine, C2-C8 carboxyl, C2-C8 ester,C1-C4 aldehyde, and glucuronide groups; L is a divalent linker includingfrom 3-7 backbone carbon atoms that form a chain connecting the Ar1,Ar2, and R11 groups as the case may be, where L includes at least one ofa carbonyl or hydroxyl group. In further embodiments, Ar1 and Ar2 arephenyl groups; R1-R10 are independently selected from the groupconsisting of H, hydroxyl, halogen, amine, nitro, sulfonate, thio,borate, C1-C2 boronate, sulfoxide, C1-C4 alkyl, C1-C4 alkoxy, C1-C4alkylamine, C2-C6 alkenylamine, C1-C6 acetoxy, C1-C4 carboxyl, with atleast one of R1-R5 and R6-R10 being hydroxyl.

b. Divalent Linking Groups

The linker L is a spacer that preferably includes 3, 4, 5, 6 or 7 carbonatoms that form a linear carbon chain connecting the first and secondaryl groups. The carbons atoms in the carbon chain that trace outshortest path between the first and optional second aryl groups arereferred to herein as the “backbone” carbon atoms. The number ofbackbone carbon atoms is readily determined in straight chain alkylgroups. In linkers that include a cyclic alkyl group as a constituent ofthe linear chain, the backbone carbon atoms include the least number ofring carbons possible. The number of backbone carbon atoms is usedherein as a shorthand way to designate the length of the linker beingused. For example, a 7-carbon linker is a divalent linker that includes7 backbone carbon atoms.

Preferably at least one of the backbone carbon atoms is included in acarbonyl (C═O) or thio carbonyl (C═S) moiety. The linker may besubstituted or unsubstituted. The linker may further be saturated orunsaturated. In a preferred embodiment, the linker contains an oddnumber of carbon atoms (i.e., 3, 5, or 7 carbon atoms), and at least oneunsaturated carbon-carbon bond. In additional embodiments, the linkermay include a hydroxyl moiety in place of, or in addition to, the atleast one carbonyl moiety.

Curcumin derivatives of the invention include a linking group L that ispreferably covalently attached at one end to aryl group Ar1. Optionally,the linking group L may also be covalently attached at the other end toa second aryl group, Ar2, which is selected independently from Ar1. Thelinking group L is a divalent linking group that preferably includes analkylene or an alkenylene group having between 3 and 7 backbone carbonatoms, and more advantageously an odd number of backbone carbon atoms(i.e., 3, 5, or 7 carbon atoms). The linker also preferably has at leastone carbonyl moiety, and may further include a hydroxyl moiety in placeof, or in addition to, the at least one carbonyl moiety. The linkinggroup may be substituted or unsubstituted, and may be saturated orunsaturated. Preferably, the linking group has a carbon-carbon doublebond between the α and β carbons relative to Ar1 and/or Ar2 (e.g., seeformulas C-1 and C-19 through C-33, which illustrate such a double bond.Still more preferably, the linking group includes conjugated doublebonds. Table 1 shows compounds with 7-carbon linkers; Table 2 showscompounds with 5-carbon linkers; and Table 3 shows compounds with3-carbon linkers.

A divalent linking group includes two carbons with unfilled valenciesthat provide valence points where a covalent bond can be formed to anadjacent alkyl or aryl group that also includes a carbon with anunfilled valency. Generally, a valence point is represented in achemical formula by a bond that is shown as not being attached toanother group (e.g., CH3-, wherein—represents the valence point).

In embodiments wherein the curcumin derivative lacks the second arylgroup Ar2, the distal valence point on the linking group can be filledwith any substituent of interest, preferably a short chain alkyl group(e.g., C1-C6, more preferably C1-C4) or a hydrogen (H). Compoundslacking a second aryl group may be represented by formula:Ar1-L-R11  C-15R11 can be, for example, a heterocyclic group or an alkyl group,preferably an alkyl group having four or fewer carbon atoms, e.g., amethyl group. R11 can alternately be an amine, a hydroxyl, a hydrogen,nitro, sulfonate, sulfoxide, thio, ester, carboxylate, amide, borate, ora C1-C4 boronate.

i. Curcumin Derivatives Including 7-Carbon Linking Groups.

In one embodiment of the invention, the curcumin derivatives include oneor two aryl groups (Ar1 and optionally Ar2) and a linking group L thatis a 7-carbon linking group (i.e., a linking group that includes 7backbone carbon atoms). Preferably, the 7-carbon linking group includesat least one unsaturated carbon-carbon bond. Examples of 7-carbonlinking groups includeCH═CH—(CO)—CR12=C(OH)—CH═CH—,  C-16CH═CH—(CO)—C(R12)2-(CO)—CH═CH—, and  C-17CH═CH—(CO)—CH═C(OH)—CH═CH—  C-18where R12 includes substituent alkyl, arylalkyl, or aryl groupscomprising 10 carbon atoms or less. In some embodiments, R12 may be amethyl, ethyl, or benzyl group. These linking groups are the divalentforms of 4-alkyl-1,6 heptadiene-3,5-dione; 4,4-dialkyl-1,6heptadiene-3,5-dione; and heptane-3,5-dione.

Table 1 shows a number of examples of curcumin derivatives that includea 7-carbon linker. The compounds shown contain two aryl rings separatedby a 7-carbon linker having two carbonyls (or the equivalent keto-enoltautomer). In many, but not all, of the compounds, the linker isunsaturated. “Bn” refers to a benzyl group.

TABLE 1 7-Carbon Linker Analogs.

C-19

C-20

C-21

C-22

C-23

C-24

C-25

C-26

C-27

C-28

C-29

C-30

C-31

C-32

C-33

C-34

C-35

C-36

C-37

C-38

C-39

C-40

C-41

ii. Curcumin Derivatives Including 5-Carbon Linking Groups

In a further embodiment of the invention, the curcumin derivativesinclude one or two aryl groups (Ar1 and optionally Ar2) that are linkedby a linking group L that is a 5-carbon linking group (i.e., a linkinggroup that includes 5 backbone carbon atoms). Preferably, the 5-carbonlinking group includes at least one unsaturated carbon-carbon bond.Examples of 5-carbon linking groups include:

These linking groups are the divalent forms of 1,4-pentadiene-3-one;pentan-3-one; pentan-3-ol, 2,6; bis(methylene)cyclohexanone; and1,2,4,5-diepoxy pentan-3-one. As noted herein, curcumin derivatives mayinclude a cyclic linking group. For example,1-methyl-2,6-diphenyl-4-piperidone provides a compound with a 5-carbonlinking group that is bridged by a tertiary amine to form a cyclicalkylene linking group including the heteroatom nitrogen.

Table 2 shows a number of examples of curcumin derivatives that includea 5-carbon linker. The compounds shown contain two aryl rings separatedby a 5-carbon linker having a single carbonyl or hydroxyl. In many, butnot all, of the compounds, the linker is unsaturated.

TABLE 2 5-Carbon Linker Analogs.

C-47

C-48

C-49

C-50

C-51

C-52

C-53

C-54

C-55

C-56

C-57

C-58

C-59

C-60

C-61

C-62

C-63

C-64

C-65

C-66

C-67

C-68

C-69

C-70

C-71

C-72

C-73

C-74

C-75

C-76

C-77

C-78

C-79

C-80

C-81

C-82

C-83

C-84

C-85

C-86

C-87

C-88

C-89

C-90

iii. Curcumin Derivatives Including 3-Carbon Linking Groups

In a further embodiment of the invention, the curcumin derivativesinclude one or two aryl groups (Ar1 and optionally Ar2) that are linkedby a linking group L that is a 3-carbon linking group (i.e., a linkinggroup that includes 3 backbone carbon atoms). Preferably, the 3-carbonlinking group includes at least one unsaturated carbon-carbon bond. Anexample of a 3-carbon linking group is —CH═CH—(CO)—; i.e., a divalentform of propenone.

Table 3 shows a number of examples of curcumin derivatives that includea 3-carbon linker. The compounds shown generally have an unsaturated3-carbon linker having a single carbonyl. While most of the examplesshown have two aryl groups separated by the linker, several of theembodiments include only a single aryl group. In the examples thatinclude only a single aryl group, a methyl group is provided at theother end of the linking group. One compound includes the heteroatom Nin place of one of the backbone carbon atoms; however, this is stillconsidered a 3-C linker in that 3 atoms (C, N, and C) are present alongthe shortest bridge between the two aryl groups.

TABLE 3 3-Carbon Linker Analogs.

C-91

C-92

C-93

C-94

C-95

C-96

C-97

C-98

C-99

C-100

C-101

C-102

C-103

iv. Additional Curcumin Derivatives

Curcumin derivatives of the invention may include a variety of linkinggroups and Ar groups while retaining the necessary activity.Accordingly, additional curcumin analogs are contemplated. These includecurcumin analogs containing central methylene substituents such asethyl, propyl, butyl, isopropyl and substituted benzyl groups accordingto the formula:

Central Methylene Substituent Analogs

Additional analogs that are contemplated are those having a pyridinering with and without a central methylene substituent on the 7-carbonlinker such as those shown in the formulas:

Pyridine Aryl Ring Analogs

Many curcumin analogs which have a 5-carbon linker possess significantactivity. Additional active analogs in this series may containsubstituents such as hydroxy and methoxy groups on the aryl rings.Examples of these analogs are shown in the formula:

Aryl Substituent Analogs

Other analogs include heterocyclic moieties, which may be substituted orunsubstituted, as shown in the formulas:

Heterocyclic Analogs

v. Synthetic Curcumin Derivatives Via Different Reaction Schemes

A large number of naturally occurring and synthetic curcuminderivatives, the latter explicated by reference to the methods ofsynthesis thereof, are disclosed in Anand et al. “Biological Activitiesof Curcumin and Its Analogues (Congeners) Made By Man and MotherNature.” Biochemical Pharmacology 76 (2008):1590-1611, which isincorporated by reference herein its entirety. Representative syntheticcurcumin derivatives are set forth below.

where R18, R19, and R20 are each independently selected from the groupconsisting of H, C1-C4 alkyl, and C1-C4 amine groups.

vi. Miscellaneous Curcumin Components

where R24 and R25 are independently selected from the group consistingof OH, C1-C4 alkoxy, and C1-C4 alkylcarbonyloxy; R26 and R27 areindependently selected from the group consisting of H, OH, C1-C4 alkoxy,and C1-C4 alkylcarbonyloxy; R28 is selected from the group consisting ofH, OH, and C1-C4 alkylcarbonyloxy; and R29 is selected from the groupconsisting of H and C1-C4 alkoxy.

where R30, R31, R32, R33, R34, R35, and R36 are independently selectedfrom the group consisting of H, OH, C1-C4 alkoxy, and C1-C4alkylcarbonyloxy.

where R37 and R38 are independently selected from the group consistingof methoxy and OH, and the center benzene ring may be substituted in the1,3- or the 1,4-position with the acryloyl groups.

Additionally, some or all of the carbonyl (C═O) moieties of formulaeC-148 through C-162 may be substituted with thio carbonyl (C═S)moieties.

vii. Presently Preferred Curcumin Components

Based upon the structure activity relationships of curcumin and curcuminderivatives, curcumin components of formula C-13A are preferred:

where R1-R5 are as previously defined, and the * denotes the valencepoint where the additional portion of the compounds are attached. Thatis, the preferred components have the moiety depicted in C-13A andadditional portion exemplified by the foregoing disclosure, e.g., theremaining structure of the linker L and Ar2 or R11.

The most preferred curcumin components are exemplified by formula C-12where:

-   -   Ar1 and Ar2 are each aryl groups, and especially phenyl,        naphthyl, thienyl, pyridinium, and pyridyl groups, and most        preferably phenyl groups, wherein all of the foregoing may be        substituted or unsubstituted;    -   L contains either 5 or 7 backbone carbon atoms and at least one        of a carbonyl or hydroxyl group;    -   at least one of R1-R5 and R6-R10 is hydroxyl.

Still more preferably, the curcumin components should have:

-   -   the R2 and R7, or R3 and R7, substituents of formulas C-13 and        C-14 as hydroxyl;    -   the R3 and R8 substituents of formulas C-13 and C-14 as methoxy        or ethoxy (most preferably methoxy);    -   a β-diketone group in the linker L of formula C-12; and/or    -   at least one, and preferably two, carbon-carbon double bonds in        the linker L of formula C-12, where at least one of the double        bonds is located between the α and β carbons relative to Ar1        and/or Ar2;    -   where the curcumin component is not apocynin.

In other embodiments, the curcumin component is taken from compounds ofthe general formulaAr1-L-Ar2  C-12where Ar1 and Ar2 are independently selected form the group consistingphenyl and naphthyl groups, where the phenyl groups may be substitutedwith one or more substituents selected from the group consisting of OH,C1-C4 alkoxys (more preferably C1-C2 alkoxys), C1-C4 haloalkyls (morepreferably C1-C2 haloalkyls), halo, and L has from 3-7 backbone carbonatoms including at least one carbonyl group therein.

Use has also been made of a commercially available, proprietary curcuminproduct sold under the designation “ResCu” by Davospharma, and which isasserted to be a more bioavailable form of standard curcumin. Thiscommercial product has been tested in connection with the invention andfound to be a suitable curcumin component.

The Harmine Component(s)

As used herein, “harmine component(s)” shall mean harmine, itsmetabolites and derivatives, isomers and tautomers thereof, and estersand pharmaceutically acceptable salts of any of the foregoing.

Harmine belongs to the family of β-carboline alkaloids, its chemicalname is 7-methoxy-1-methyl 9H-pyrrole[3,4-b]indole, and its molecularformula is C13H12N2O. The base structure of β-carboline is shown in H-0.

Harmine has a molecular weight of 212.25 and a melting point of 261° C.Harmine was originally isolated from Peganum harmala, which is widelyused as a traditional herbal drug in the Middle East and North Africa.The chemical structure of harmine, 1-methyl-7-methoxy-β-carboline, isshown as follows:

Harmine derivatives, which are variously substituted β-carbolines, areillustrated in the following formulas, which differ in that H-2 has aquaternary ammonium group at the 2-position, whereas H-3 has a tertiarynitrogen at the 2-position.

With reference to formulas H-2 and H-3:

-   -   Z1 is hydrogen; a C1-C6 alkyl or haloalkyl, a C2-C6 alkenyl or        haloalkenyl; an aryl group or an arylalkyl group, wherein the        aryl group is optionally substituted at any position with        halogen, nitro, hydroxyl, C1-C3 alkoxy, amino, sulfonate,        sulfoxide, thio, ester, carboxylate, amide, borate, or C1-C4        boronate and wherein the alkyl group is selected from C1-C4        alkyl; or a heterocyclic group;    -   Z2 is hydrogen; a C1-C6 carboxyl, ester, carboxylate, acylamino,        acyl halide, sulfonate, sulfoxide, thio, amide or alkoxycarbonyl        group; an aryloxycarbonyl group; alkyl group optionally        substituted with hydroxyl or alkoxycarbonyl; carbamate;        acylhydrazine; or a heterocyclic oxycarbonyl group, where the        heterocyclic portion contains from 3-7 atoms and a nitrogen,        oxygen, boron, or sulfur heteroatom;    -   Z3 is hydrogen; a C1-C6 alkyl or haloalkyl; sulfonate,        sulfoxide, thio, carboxylate, amide, C2-C6 alkenyl group;        hydroxyl; a C1-C6 alkoxy group; a C1-C6 carboxylic ester group;        an arylalkoxy group where the alkoxy portion contains from 1-6        carbon atoms; or a heterocyclic group containing from 3-7 atoms        and a nitrogen, oxygen, boron, or sulfur heteroatom;    -   Z4 is hydrogen; a C1-C6 alkyl, haloalkyl; C2-C6 alkenyl group; a        hydroxyalkyl group where the alkyl portion contains from 1-6        carbon atoms; an arylalkyl group wherein the aryl group is        optionally substituted at any position with halogen, nitro,        hydroxyl, C1-C3 alkoxy, borate, C1-C4 boronate, sulfonate,        sulfoxide, thio, ester, carboxylate, amide or amino, and wherein        the alkyl group is selected from C1-C4 alkyl group; an        arylalkanone; or a heterocyclic group containing from 3-7 atoms        and a nitrogen, oxygen, boron or sulfur heteroatom;    -   Z5 is hydrogen; a C1-C6 alkyl group; an aryl group substituted        at any position with one or more of (1)-(2), where (1) is a        C1-C4 alkyl group, and (2) is a C1-C6 carbonyl, hydroxycarbonyl,        ester, sulfonate, sulfoxide, thio, ester, carboxylate, amide or        amino group; arylalkyl where the alkyl group is a C1-C6 alkyl;        1-5 substituted arylalkyl; arylhydrocarbyl; arylcarboxyl; aryl        ester group; arylamino group; or a heterocyclic group containing        from 3-7 atoms and a nitrogen, oxygen, boron or sulfur        heteroatom;    -   X is a halogen; a sulfonic group, a sulfuric group, a nitric        acid group or a carboxylate, in the compounds of the above        formulas H-2 and H-3:    -   Z1 is preferably hydrogen, a C1-C4 alkyl group, or an arylalkyl        group; more preferably hydrogen, or a C1-C2 alkyl group; and        most preferably methyl;    -   Z2 is preferably hydrogen or a C1-C4 alkoxycarbonyl group; more        preferably a C1-C2 alkoxycarbonyl group; and most preferably        hydrogen;    -   Z3 is preferably hydrogen, hydroxyl, or a C1-C4 alkyloxy group;        more preferably methoxy;    -   Z4 is preferably hydrogen, a C1-C4 alkyl group, a C1-C4        hydroxyalkyl group, or an optionally substituted arylalkyl        group; more preferably hydrogen, a C1-C2 alkyl group, or a C1-C2        hydroxyalkyl group; still more preferably ethyl or benzyl; and        most preferably hydrogen.

In certain embodiments where Z1 is hydrogen, a C1-C4 alkyl group, or anarylalkyl group, Z2 is hydrogen, hydroxyl, a C1-C4 carboxyl group, aC1-C4 ester group, a carboxylate group, a halogen, or a C1-C4alkoxycarbonyl group; Z3 is hydrogen, hydroxyl, or a C1-4 alkoxy group;Z4 is hydrogen, a C1-C2 alkyl group, a C1-C2 hydroxyalkyl group, or anoptionally substituted arylalkyl group; and Z5 is hydrogen, a C1-C6alkyl group, or an optionally substituted aryl-alkyl group.

In certain embodiments where Z1 is hydrogen, Z2 is a C1-C2alkoxycarbonyl group, Z3 is hydrogen, and Z4 is C1-C2 alkyl group, or anoptionally substituted arylalkyl group.

In certain embodiments where Z1 is hydrogen, Z2 is ethoxycarbonyl, Z3 ishydrogen, and Z4 is ethyl or benzyl.

In certain embodiments where Z1 is methyl, Z2 is ethoxycarbonyl, Z3 ishydrogen, Z4 is pentafluorobenzyl, and Z5 is hydrogen.

In certain embodiments where Z1 is hydrogen, Z2 is hydrogen, Z3 ishydrogen, Z4 is benzyl, Z5 is benzyl, and X is bromine.

Certain preferred harmine components include β-carboline, tryptoline,pinoline, harmine, harmalol, harmalol hydrochloride hydrate,tetrahydroharmine, harmane, harmol, vasicine, and vasicinone.

Other harmine derivatives include:

where Z1, Z2, and Z9 are set forth below, and X1 is a halogen preferablyCl, Br, and F or carboxylate anion.

Based upon the structure-activity relationships of harmine and harminederivatives, preferred components are illustrated by formula H-6:

where the three * denote valence points where respective additionalatoms or portions of the components are attached to give completecompounds; the dotted lines in the right-hand six-membered ring of H-6indicate that the ring may optionally be aromatic; Z1 and Z4 areindependently H or a C1-C6 alkyl group, more preferably a C1-C4 alkylgroup, and most preferably methyl for Z1 and hydrogen for Z4; and theremaining unoccupied positions on the six-membered rings (i.e., theleft-hand phenyl group and the right-hand six-membered ring) are eachindependently selected from the group consisting of Z1, as previouslydefined. That is, the preferred harmine components have the moietydepicted in H-1 and additional portions exemplified by the foregoingdisclosure. N+ is optionally substituted with Z5, as previously defined.In some cases, X⁻ as previously defined is present. One of ordinaryskill in the art would understand that

indicates a double or single bond is present. For example, where one ofthe -* substituents is OCH3 and is located at the No. 7 position asillustrated in formula H-1, the other -* substituents on the remainingunoccupied positions on the terminal 6-membered rings are all H, Z4 is Hand Z1 is methyl, and Z5 and X⁻ not existing, the resulting compound isharmine.

Z6 Z7 Z8

CH₃CH₂CH₂CH₂—

CH₃—

CH₃CH₂CH₂CH₂—

CH₃CH₂CH₂CH₂— H—

CH₃CH₂CH₂CH₂—

CH₃CH₂CH₂CH₂—

CH₃CH₂CH₂CH₂—

H— CH₃CH₂CH₂CH₂—

One class of preferred harmine components are shown in formula H-3,where Z3 is a methoxy or ethoxy group and Z4 is a benzyl group. A secondpreferred class of harmine components is shown in formula H-2, where Z1is a methyl group, Z2 is hydrogen, Z3 is a benzyloxy group, Z4 and Z5are benzyl groups.

In other embodiments, preferred harmine components include compounds ofthe formula

R39 is selected from the group consisting of H, C1-C4 alkoxys (morepreferably C1-C2 alkoxys), O, and OH substituents, R40 is selected fromthe group consisting of H, C1-C4 alkyls (more preferably C1-C2 alkyls),C1-C4 haloalkyls (more preferably C1-C2 haloalkyls), C1-C4 alkoxys (morepreferably C1-C2 alkoxys), nitrophenyls, C1-C4 organic acids (morepreferably C1-C2 organic acids), C1-C4 alkylalcohols (more preferablyC1-C2 alkylalcohols), and C1-C6 alkyl esters (more preferably C2-C4alkyl esters), and R41 is selected from the group consisting of nothing,H, and nitrophenyls.The Isovanillin Component(s)

As used herein, “isovanillin component(s)” refers to isovanillin, itsmetabolites and derivatives, isomers and tautomers thereof, and estersand pharmaceutically acceptable salts of any of the foregoing.

Isovanillin is a phenolic aldehyde having a hydroxyl group at the metaposition and a methoxy group at the para position. Isovanillin isillustrated in the following structure:

Isovanillin is metabolized in vivo to vanillin, which is the same asstructure I-1, except that the hydroxyl and methoxy substituents areexchanged, i.e., in vanillin, the hydroxyl group is in the paraposition, and the methoxy group is in the meta position.

Useful derivatives of isovanillin have the following general formula:

where at least one of Q1-Q6 is an alkoxy group and/or an aldehyde group,especially where the alkoxy and/or aldehyde groups contain from 1-6carbon atoms; more preferably, where Q1 is an aldehyde, alcohol, amine,carbonyl, carboxylate, C1-C6 alkylhydroxy, ester, imidazole, nitro,sulfonate, sulfoxide, thio, amide, oxime, borate, or boronate, orsemicarbazone group; the Q2-Q6 groups are independently selected fromhydrogen, hydroxyl, halo, nitro, C1-C6 alkoxy, C1-C6 alkyl or alkenylgroups, with the proviso that at least one of the Q2-Q6 groups is analkoxy group. In more preferred forms, the aldehyde, alcohol, amine,carbonyl, carboxylate, and ester groups should have a C1-C6 carbon chainlength, the boronate is a C1-C4 boronate, and at least one of the Q2-Q6groups is an alkoxy group (most preferably methoxy), and another is ahydroxyl group; advantageously, the alkoxy and hydroxyl groups areadjacent each other. In particularly preferred forms, the remainder ofthe Q2-Q6 groups, apart from the alkoxy and hydroxyl groups, are all H.Advantageously, the isovanillin components of the invention shouldinclude only one phenyl group and are free of any fused ring structures(as used herein, “fused ring structures” refer to structures such asfound in naphthalene or anthracene where two rings share common atoms).Some preferred isovanillin components are selected from isovanillin,vanillin, ethyl vanillin, ortho-vanillin, vanillic acid, isovanillicacid, vanillic alcohol, isovanillic alcohol,6-bromine-5-hydroxy-4-methoxybenzaldehyde,4-hydroxy-3,5-dimethoxybenzaldehyde,4,5-dihydroxy-3-methoxybenzaldehyde, 5-hydroxy-4-methoxybenazldehyde,2-Benzyloxy-3-methoxybenzaldehyde,2-(2-benzyloxy-3-methoxyphenyl)-1H-benzimidazole,N-1-(2-benzyloxy-3-methoxybenzyl)-2-(2-benzyloxy-3-methoxyphenyl)-1H-benzimidazole,and (S)-1-(2-benzyloxy-3-methoxyphenyl)-2,2,2-trichloroethylbenzenesulfonate (regarding the last four compounds, see Al-Mudaris etal., Anticancer Properties of Novel Synthetic Vanillin Derivatives(2012)).

Certain imidazole derivatives (such as nitro-imidazoles) have been shownto have significant anticancer activity, see, e.g., Sharma et al.,“Imidazole Derivatives Show Anticancer Potential by Inducing Apoptosisand Cellular Senescence,” Med. Chem. Commun. 2014, 5, 1751, incorporatedby reference herein. Representative imidazole derivatives include:

Additional isovanillin derivatives of interest are:

It will be appreciated that the aryl group substituents in formulasI-14-I18 can be at any desired position on the on the aryl rings, e.g.,the —OH need not be adjacent and the heteroatom substituents may be atany desired location.

Based upon the structure-activity relationships of isovanillin andisovanillin derivatives, preferred components are illustrated by formulaI-13:

where the two * denote valence points where respective additional atomsor portions of the components are attached to form complete compounds,the remaining phenyl ring positions not taken by the two depictedmoieties are independently R1 as previously defined with reference toformula C-13, and Q7 is H or a C1-C6 alkyl group, more preferably aC1-C4 alkyl group, and most preferably methyl. That is, the preferredisovanillin components have the moiety depicted in 1-13 and additionalportions exemplified by the foregoing disclosure. For example, where *is CHO, the substituent —O-Q7 is located at the para position, Q7 is amethyl group, and the —O—* group is located at the meta position, the *of the —O—* moiety is H, and the remaining unoccupied positions on thephenyl ring are all H, the resulting compound is isovanillin. Inpreferred forms, the isovanillin component is not apocynin.

In other embodiments, the isovanillin components are selected from thecompounds of the formula

where Q9 is selected from the group consisting of C1-C4 aldehydes (morepreferably C1-C2 aldehydes), C1-C4 alkylalcohols (more preferably C1-C2alkylalcohols), C1-C4 alkyl esters (more preferably C1-C2 alkyl esters),and C1-C4 organic acids (more preferably C1-C2 organic acids), Q10 isselected from the group consisting of OH, H, C1-C6 alkyl esters (morepreferably C1-C4 alkyl esters), C1-C4 alkoxys (more preferably C1-C2alkoxys), and benzyloxy, and Q11 is selected from the group consistingof H, C1-C4 alkoxys (more preferably C1-C2 alkoxys), and OH.The Complete Compositions of the Invention

In the case of the preferred three-component compositions of theinvention, made by combining individual quantities of normally highlypurified curcumin component(s), harmine component(s), and isovanillincomponent(s), the as-added amounts should give weight ratios of about10:1.7:0.85 (isovanillin component(s):harmine component(s):curcumincomponent(s)), but more broadly, the ratios are approximately0.1-25:0.1-5:0.1-5 (isovanillin component(s):harminecomponent(s):curcumin component(s)). In this respect, it will be seenthat the isovanillin component(s) of the preferred GZ17-6.02 productis/are the preponderant component(s) in the compositions on a weightbasis, with the harmine and curcumin component(s) being present inlesser amounts on a weight basis. Generally, the isovanillincomponent(s) in the most preferred product should be present at a levelat least three times (more preferably at least five times) greater thanthat of each of the harmine and curcumin component(s), again on a weightbasis. However, the invention is not limited to such weight ratios. Asnoted in Example 17, an isovanillin component:harmine component:curcumincomponent weight ratio of 1:1:1 is also effective. In terms of amountsof the three components, the isovanillin component(s) should be presentat a level of from about 25-85% by weight, the harmine component(s)should be present at a level of from about 7-50% by weight, and thecurcumin component(s) should be present at a level of from about 5-40%by weight, based upon the total weight of the three components taken as100% by weight.

In the case of two-component compositions in accordance with theinvention made up of curcumin component(s) plus harmine component(s),the weight ratio of the curcumin component(s):harmine component(s)should range from about 0.01:1 to 10:1; and the weight amounts of thecurcumin component(s) should range from about 20% to 75% by weight (morepreferably from about 30-55% by weight, and most preferably about 45% byweight), and the weight amounts of the harmine component(s) should rangefrom about 25% to 80% by weight (more preferably from about 45% to 70%by weight, and most preferably about 55% by weight), based upon thetotal weight of the two components in the compositions taken as 100% byweight. Generally, the amount of the harmine component should be greaterthan the amount of the curcumin component in these two-componentcompositions.

Two-component compositions made up of isovanillin component(s) pluscurcumin component(s) should have a weight ratio of the isovanillincomponent(s):curcumin component(s) ranging from about 0.5:1 to 25:1; andthe weight amounts of the isovanillin component(s) should range fromabout 25% to 95% by weight (more preferably from about 75% to 95% byweight, and most preferably about 88% by weight), and the weight amountsof the curcumin component(s) should range from about 5% to 75% by weight(more preferably from about 5% to 25% by weight, and most preferablyabout 12% by weight), based upon the total weight of the two componentsin the compositions taken as 100% by weight.

Finally, two-component compositions made up of isovanillin component(s)plus harmine component(s) should have a weight ratio of the isovanillincomponent(s):harmine component(s) should range from about 0.5:1 to 15:1;and the weight amounts of the isovanillin component(s) should range fromabout 25% to 95% by weight (more preferably from about 75% to 95% byweight, and most preferably about 85% by weight), and the weight amountsof the harmine component(s) should range from about 5% to 75% by weight(more preferably from about 5% to 25% by weight, and most preferablyabout 15% by weight) by weight, based upon the total weight of the twocomponents in the compositions taken as 100% by weight.

In this connection, it should be understood that the isovanillincomponent(s) of any of the foregoing two- or three-componentcompositions containing isovanillin component(s) may be made up ofinitially added isovanillin component(s) plus any degradation productsof the other component(s) yielding products within the ambit of theisovanillin component(s); for example, it is known that under certaincircumstances curcumin will spontaneously degrade to give quantities ofvanillin, and, in such circumstances, the total amount of theisovanillin component(s) would be the initially added amounts togetherwith these degradation products. More broadly, the ultimate amounts ofthe curcumin, harmine, and isovanillin component(s) in the compositionsof the invention should be determined based upon the actual contents ofthe component(s) in question, regardless of whether these component(s)are derived from the originally added component(s) or as degradationproducts of some or all of these originally added component(s).

As indicated previously, levels of dosing using the compositions of theinvention is quite variable owing to factors such as the patient's age,patient's physical condition, the type of condition(s) being treated(e.g., specific cancer(s)), and the severity of the conditions. Ingeneral, however, regardless of the dosage form or route ofadministration employed, such as liquid solutions or suspensions,capsules, pills, or tablets, via oral, parenteral, or injection, thecompositions should be dosed of from about 5 to 2000 mg per day, andmore usually from about 100-800 mg per day. Such dosages may be based ona single administration per day, but more usually multipleadministrations per day.

Finally, the invention also embraces compositions and methods where theindividual components are provided and administered separately tosubjects, so long as the therapeutic effects of the invention aresubstantially preserved.

Examples 1-28

The following Examples set forth preferred therapeutic agents andmethods in accordance with the invention, but it is to be understoodthat these examples are given by way of illustration only, and nothingtherein should be taken as a limitation upon the overall scope of theinvention. A number of the Examples set forth various tests using thepreferred drug of the invention, GZ17-6.02, which is sometimes referredto as GZ17Syn-6.02.

The GZ17-6.02 product of the Examples was made by dispersing quantitiesof solid synthetic isovanillin (771 mg, 98% by weight purity), syntheticharmine (130.3 mg, 99% by weight purity), and a commercially availablecurcumin product derived by the treatment of turmeric (98.7 mg,containing 99.76% by weight curcuminoids, namely 71.38% curcumin, 15.68%demethoxycurcumin, and 12.70% bisdemethoxycurcumin), in a 1 mL ethanolat a weight ratio of 771:130.3:98.7 (isovanillin:harmine:curcuminproduct) in ethanol followed by sonication of the dispersion. Aliquotsof this stock solution were then used to create the different GZ17-6.02dilutions using stock media of the cells in question.

The two-component products described in Example 23 were made in the samefashion as the GZ17-6.02 agent, and the weight ratios of the twocomponents were as set forth immediately above, e.g., theisovanillin/harmine sub-combinations contained a weight ratio of771:130.3, and so forth.

Example 1

In this example, the preferred GZ17-6.02 product was tested with twodifferent human head and neck cancers (HN5 and OSC19), in order todetermine the extent of cell death induced by the product.

Methods

The respective cells were individually cultured in a growth mediumprepared using RPMI-1640 medium containing 11.1 mM D-glucose with 10%fetal bovine serum, 10 mM HEPES, 2 mM L-glutamine, 1 mM sodium pyruvate,0.05 mM 2-mercaptoethanol, and Antibiotic-Antimycotic. These cells weremaintained in T75 tissue culture flasks in a humidified incubator at 37°C. and 5% CO₂. The media were changed on a third day after cell plating,and the cells were passaged on day 5 by trypsinization.

Formation of Cancer Spheroids

Custom-made micromolds with 100 μm diameter wells were loaded with thecells (U.S. Pat. No. 8,735,154, incorporated by reference herein). Themedia were changed every day by partial replacement. Cell aggregateswere allowed to form in the micromolds for 72 hours and then weretransferred to a 100 mm non-treated tissue culture dishes for 3additional days. The spheroids were passed through 70 μm and 100 μm cellstrainers (#3431751 and #43152, Corning, Tewksbury, Mass.) andmaintained in HEPES Balanced Salt Solution comprised of: 20 mM HEPES,114 mM NaCl, 4.7 mM KCl, 1.2 mM KH₂PO₄, 1.16 mM MgSO₄, 2.5 mM CaCl₂),25.5 mMNaHCO₃ and 0.2% bovine serum albumin (fatty acid free), pH 7.2.

Testing of GZ17-6.02 on Head and Neck Cancer Spheroids

Individual wells of a 96 well plate were manually loaded with 15-20cancer spheroids each, and exposed to selected doses of GZ17-6.02. Eachtrial was run with at least 4 replicates at each dose. After a 24 hourexposure to the selected dosages of GZ17-6.02, PrestoBlue (LifeTechnologies, Inc) was added to each well and fluorescence readings weretaken 4-6 hours later with an excitation wavelength of 485 nm and anemission wavelength of 560 nm, using a microplate reader (EnspireMultimode, PerkinElmer). Results were averaged following backgroundsubtraction and normalized to untreated cell controls.

FIG. 1 demonstrated that GZ17-6.02 dramatically kills HN5 and OSC19cancer cells in a dose-dependent manner. The X axis is the increasingdose of GZ17-6.02 and the Y axis is the average number of live cells ineach well at the end of a 24 hour exposure to the stated dose ofGZ17-6.02.

Example 2

In this example, GZ17-6.02 was found to induce significant cancer celldeath in human pediatric leukemia cells and pediatric osteosarcoma in adose-dependent manner.

Jurkat leukemia cells were grown in suspension in media (RPMIsupplemented with 10% FBS), maintained at approximately 500,000cells/mL. The cells were plated in 96-well plates, and each well wasexposed to a selected dose of GZ17-6.02 for 24 hours (a minimum of 4replicates for each dosage). These cells were not treated to generatespheroids, but were directly plated onto the well plates. After a 24hour exposure to the selected dosages of GZ17-6.02, PrestoBlue (LifeTechnologies, Inc) was added to each well and fluorescence readings weretaken 4-6 hours later with an excitation wavelength of 485 nm and anemission wavelength of 560 nm, using a microplate reader (EnspireMultimode, PerkinElmer). Results were averaged following backgroundsubtraction and normalized to untreated cell controls.

Human osteosarcoma cells (HOS) also had a significant dose-dependentincrease in cell death when exposed to GZ17-6.02 for 24 hours. Themethods to create and test osteosarcoma spheroids were the same as themethods described in Example 1.

As illustrated in FIGS. 2A and 2B, increasing doses of GZ17-6.02 inducedsignificant cell death in both leukemia and osteosarcoma cell typestested.

Example 3

In this example, the effect of increasing doses of GZ17-6.02 in killinghuman lymphoma cells (mo205) and human lung cancer cells (H358) wastested.

The lymphomas treatment methods used were identical to those describedin Example 2 relative to the pediatric leukemia cells, whereas the lungcancer treatment method was the same as described in Example 1.

FIGS. 3A and 3B set forth the results of these tests, and demonstratethe effectiveness of GZ17-6.02 in inducing lymphoma and lung cancer celldeath.

Example 4

In this example, the effect of increasing doses of GZ17-6.02 in killinghuman ovarian cancer cells (A1847) and human prostate cancer cells(22rv1) was tested.

The cells were treated and tested as set forth in Example 1.

This test confirmed that both types of cells experienced dose-dependentdeath when exposed to GZ17-6.02, see FIGS. 4A and 4B.

Example 5

In this example, the effect of increasing doses of GZ17-6.02 in killinghuman breast cancer cells (du4475) and human pancreatic cancer cells(panc-1) was tested.

The cells were treated and tested as set forth in Example 1, except thata different growth medium was used, namely Dulbecco's Modified EagleMedium (DMEM) with 10% fetal bovine serum, and 1%penicillin/streptomycin mix (Sigma-Aldrich).

This test confirmed that both types of cells experienced dose-dependentdeath when exposed to GZ17-6.02, see FIGS. 5A and 5B.

As is evident from Examples 1-5, ten different cancer types were testedwith GZ17-6.02 and they all were sensitive to the therapeutic agent. Inmost of the cancers (head and neck cancer (OSC19), leukemia,osteosarcoma (bone cancer), lymphoma, lung cancer, ovarian, prostate,breast, and pancreatic cancer), the compound killed nearly all of thecells in the dish at a dose of 3.13-6.25 mg/mL. This array of cancertypes represents a substantial portion of human cancers. Moreover, theresults confirm that the therapeutic agent is effective in both solidtumor cancers and blood cancers.

Example 6

In this test, non-cancerous integumental (dermal) fibroblasts (hgf-1)were tested with GZ17-6.02 and compared with prostate cancer (22rv1) andovarian cancer cells (A1847), to determine the effect of GZ17-6.02 onthe non-cancerous cells versus the prostate cancer and ovarian cancercells.

The fibroblasts were treated as follows: The cells were grown toconfluence in the DMEM medium of Example 5, and then placed in 96-wellplates where they adhered to the bottom of the plates. Each well wasthen exposed to a selected dose of GZ17-6.02 for 24 hours (a minimum of4 replicates each), and tested according to Example 1. The prostatecancer and ovarian cancer results were taken from the previous Example4.

As illustrated in FIG. 6, the cancer cells begin to die at lowerGZ17-6.02 doses than the non-cancerous fibroblast cells, demonstratingthat GZ17-6.02 is more toxic to cancer cells versus the non-cancerousfibroblasts.

Example 7

Migration of cells away from a primary tumor, followed by invasion ofthose tumor cells first into the blood vessels and later out of theblood vessels and into other tissues, are the two steps essential formetastases to form from a primary tumor. The cells were grown in theDMEM media of Example 5.

Head and neck cancer cells (OSC19) were tested using a commerciallyavailable migration assay kit, Cultrex 96 Well Migration Assay (R&DSystems). The cell movement from the upper compartment to the lowercompartment was measured as representative of cell migration. A negativecontrol migration was also measured, using the GZ17-6.02 vehicle, namelya dilution of ethanol in the growth media to the same concentration asGZ17-6.02 in the growth media.

The head and neck cancer cells were also assayed for migration using aCultrex Cell Invasion Assay (R&D Systems). The ability of the head andneck cancer cells to invade a collagen matrix was measured by countingcells that exited the upper chamber and passed through acollagen-surrounded lower chamber. Similarly, a negative control forinvasion was also measured, using the same ethanol vehicle employed inthe migration assay.

As illustrated in FIG. 7A, the GZ17-6.02 significantly inhibitedmigration so that almost no head and neck cancer cells could migratefrom the site of origin. Likewise, GZ17-6.02 significantly inhibitedinvasion (FIG. 7B) by more than 60%.

Example 8

In this test, the synergism of GZ17-6.02 with doxorubicin (a commonchemotherapy drug) was tested on ovarian cancer cells. In this test,increasing doses of doxorubicin were administered to ovarian cancerspheroids and cell deaths were measured, as described in Example 1. Theprotocol involved testing doxorubicin alone at various dosages (FIG. 8,doxorubicin alone—closed circles), with comparative tests using thevarious dosages of doxorubicin in combination with either 0.2 mg/mL(FIG. 8 open circles) or 0.4 mg/mL (FIG. 8 closed triangles) GZ17-6.02.Additionally, the induced cell death results from use of 0.2 mg/mLGZ17-6.02 alone (FIG. 8, line A) and 0.4 mg/mL GZ17-6.02 alone (FIG. 8,line B) were determined.

As is evident from FIG. 8, even at the lowest dose of doxorubicin (0.002the addition of GZ17-6.02 at 0.2 and 0.4 mg/mL increased cell deathsignificantly below the level of either compound alone. At low doses ofdoxorubicin alone (up to 0.158 doxorubicin was ineffective. GZ17-6.02administered alone at a dose of 0.2 mg/mL resulted in cell death at alevel represented by the upper horizontal dotted line in FIG. 8.Moreover, GZ17-6.02 administered alone at a dose of 0.4 mg/mL resultedin cell death at a level represented by the lower horizontal dotted linein FIG. 8. Thus, even at these low dosages, addition of GZ17-6.02significantly increased cell death (see data points under both of thehorizontal dotted lines in FIG. 8). This demonstrates the synergy of thecombination of GZ17-6.02 and doxorubicin, even at low dosages of bothGZ17-6.02 and doxorubicin. This trend is confirmed at higher dosages ofdoxorubicin, where the combined products were always more effective thandoxorubicin alone.

Example 9

There are multiple general ways in which cancer (blood or solid tumors)can be blocked by chemotherapy. One is to actually induce death in thecancer cells (apoptosis), and a second is to block the ability of cellsto receive nutrients, basically starving them to death (necrosis). Thistest demonstrates that at the ED₅₀ of GZ17-6.02 on head and neck cancercells (OSC19) caused cell death via apoptosis. In the test, head andneck cancer cells were stained with an apoptotic fluorophore and sortedby flow cytometry, which allows a single cell to be counted and measuredfor fluorescence simultaneously. This histograms in FIGS. 9A and 9Billustrate the shift in fluorescence intensity to a lower level,indicating apoptotic cell death. While the control (FIG. 9A, head andneck cancer cells without GZ17-6.02) had little apoptotic cell death(19.8%), 48.2% of the cells exposed to GZ17-6.02 died of apoptosis (FIG.9B). This indicates one type of mechanism of action of GZ17-6.02.

Example 10

Caspase proteins are intracellular proteins involved in the apoptoticprogrammed cell death pathway. In instances where live lung cancer cellnumber decrease while a caspase protein increases, it can be inferred byassociation that the caspase was activated, and was thus associated withthe molecular pathway of the cell death.

In these tests, the lung cancer cells (H358) were assayed using caspasetesting kits (Promega Caspase-Glo 3/7, -Glo 2, -Glo 6, -Glo 8, and -Glo9). Sample lung cancer cells were homogenized on ice in the providedpreparation buffer and agitated on a plate shaker at low speeds (around300 rpm). The supernatants were exposed to the provided assay solutionsand read on a microplate reader for luminescent output (EnspireMultimode, PerkinElmer).

The test results indicated that caspases 3 and 7 (FIG. 10A, filledtriangles) increased dramatically in association with cell death (filledcircles). In addition, results indicated that caspase 6 (FIG. 10B, opencircles) increased in association with cell death (filled circles). Incontrast, caspase 9 was not activated (FIG. 10A, open circles). Thisimplies a receptor-mediated cell death pathway and not a mitochondrialcell death pathway. In order to confirm this, an ATP assay(CellTiter-Glo, Promega) was conducted in which toxicity of themitochondria were measured at the same time as cell death, based uponATP concentrations. If a mitochondrial toxicity were responsible forcell death, it would happen at lower doses or prior in time to the celldeath. FIG. 10C confirms that there was no significant mitochondrialtoxicity (filled circles) while there was cell death (open circles).

Example 11

The procedures of Example 10 were followed, with testing of GZ17-6.02 onovarian cancer cells (A1847). Caspases 3 and 7 were activated at lowdoses of GZ17-6.02, indicating a receptor-mediated cell death (FIG. 11A,filled circles). Caspase 6 was active at low doses of GZ17-6.02 (FIG.11B, filled circles). However, caspase 8 was activated, which typicallysignals cell death via the mitochondria (FIG. 11C, filled circles) andcaspase 9 was also activated (FIG. 11D, filled circles). In order toconfirm the mitochondrial cell death pathway, mitochondrial toxicity wasmonitored in comparison to cell death (FIG. 11E, filled circles),indicating that the mitochondrial toxicity was at least partiallyinvolved in the subsequent cell death.

Example 12

The procedures of Example 10 were followed, with testing of GZ17-6.02 onosteosarcoma cells (HOS). These tests indicated that levels of caspases3 and 7 were significantly higher when exposed to doses of GZ17-6.02over 0.4 mg/mL before or during cell death (FIG. 12A, filled circles).Caspase 6 was activated at high doses of GZ17-6.02 (FIG. 12B, filledcircles). In contrast, caspase 2 (FIG. 12C, filled circles) and caspase9 (FIG. 12D, filled circles) showed no activation that would indicatethat they were involved in the cell death pathways induced by GZ17-6.02.

Example 13

The procedures of Example 10 were followed, with testing of GZ17-6.02 onhuman head and neck cancer cells (OSC19). FIG. 13 (filled circles)indicates that there is no mitochondrial toxicity involved in GZ17-6.02induced cell death in the head and neck cancer cells.

Example 14

In addition to directly killing cancer cells, chemotherapeutic agentscan work by blocking the rapid proliferation of cancer cells, thuseventually halting the progression of the cancer.

In this test, a multiplex dot-based Western assay for proliferativeproteins was used to measure the relative amounts of proteins known tobe involved in cell proliferation signaling, in both a control test (noGZ17-6.02) and an inactivation test (with GZ17-6.02, at the ED₅₀ forhead and neck cancer cells).

Following a 24 hour exposure to GZ17-6.02 or vehicle head and neckcancer cells (OSC19) were homogenized and the lysate loaded onto theHuman Phospho-Kinase Array Kit (R&D Systems) using the followingprocedure. The antibody-attached membranes were incubated overnight at4° C. with the respective cell lysates, followed by repeated washing andincubation in the detection antibodies for 24 hours. After subsequentrepeated washes, streptavidin-HRP was applied for 30 minutes. Membraneswere washed and exposed to film after applying chemiluminescence reagentmix.

Cellular proteins specific to a pathway of cell proliferation weretested in duplicate (resulting in 2 side-by-side dots for each proteinin FIG. 14A). The darker the dot, the more relative protein was presentin the homogenate. As shown in FIG. 14B, cell proliferation proteinsshowed a dramatic reduction in amount when exposed to the head and neckcancer cell ED₅₀ dose of GZ17-6.02. The proteins showing the greatestreduction included epidermal growth factor receptor (block a compared toblock a′), extracellular-signal-regulated kinase (block b compared toblock b′), the catalytic subunit of AMP-activated kinase (block ccompared to block c′), β-catenin (block d compared to block d′), STAT2(block e compared to block e′) and Chk-2 (block f compared to block f).

Example 15

In this example, two independent scientists conducted identical testswith ovarian cancer and lung cancer cells (A1847 and H358) using thetechniques described in Example 1, in order to determine induced celldeath upon application of GZ17-6.02. The nearly identical results(filled circles versus open circles) confirm the reliability of thetesting as a determination of GZ17-6.02 effectiveness. FIG. 15A setsforth the ovarian cancer results, whereas FIG. 15B gives the lung cancercell results.

Example 16

Six-week old FOXN1 mice were inoculated with human head and neck cancercells (OSC19) bilaterally into the flank region, in order to induce theformation of palpable tumors. The bilateral tumors were measured at oneweek after cell injection, and had each grown to an average volume ofapproximately 9 mm³. Half of the mice were injected daily with 15 mg/kgbody weight GZ17-6.02 stock solution into the right side tumors,starting on day 7 after inoculation of the tumor cells. The other halfof the mice were injected with the same volume of the ethanol carrier ofthe stock solution into the right side tumors. Thus, each mouse had 2palpable tumors, one on either flank, but only one side was injectedwith GZ17-6.02. Every 2-4 days, the tumors on both sides of each mousewere measured with vernier calipers in 2 perpendicular dimensions andthe volumes calculated.

FIG. 16A shows that GZ17-6.02 dramatically halted tumor growth in thetreated mice, so that by 3 weeks there was a distinct and statisticallysignificant difference between the vehicle-treated controls and theGZ17-6.02 treated mice. Further, the trend in the GZ17-6.02 treated micewas that the tumors began to decrease in volume from day 21 (althoughthe decrease was not statistically significant) from tumor size at day28. FIG. 16A shows the decline in tumor volume in the tumors that weredirectly injected with GZ17-6.02. FIG. 16B shows the halt in tumorgrowth that occurred in the non-injected tumors on the contralateralside of the neck of the treated mice. FIG. 16B demonstrates thatGZ17-6.02 has a systemic anticancer effect, i.e., the contralateraltumor size decrease without direct injection of GZ17-6.02 indicates thatthe agent traveled through the bloodstream of the mice.

Throughout the study, the animals showed no signs of complications,distress, or toxicity. Thus, none of the over 30 mice in the studyexhibited any observable adverse reactions. Daily observation of themice documented that none suffered any weight loss, new tumor formation,loss of appetite, change in fur appearance or grooming behavior, orchange in activity owing to lethargy. Moreover, there were no grossabnormalities of any of the internal organs of the mice upon necropsy.It was thus concluded that there were no drug-drug interactions, whichare common with multiple-drug anti-cancer compositions.

In another mice study, a head and neck tumor was surgically removed froma human patient, and approximate 35 mg-portions of the tumor wereimplanted in a first randomized group of ten nude-FOXN1 mice using a 5%ethanol in saline vehicle via oral gavage. A second randomized group often mice was treated only with the vehicle as a control, in the samemanner as the first group. The first mice group was treated with 30mg/kg/day doses of GZ17-6.02 five days/week, and the doses wereincreased to 50 mg/kg/day during the second week of treatment.

The two groups of mice were treated for a total of three weeks, andtumor volumes were measured twice a week using a Vernier caliper. Theresults of this study are set forth in FIG. 16C, which illustrates thatthe fractional tumor volumes relative to the pretreatment tumor sizewere reduced in the first group of mice, while the control miceexhibited gradually increasing tumor burdens.

Example 17

In this Example, a different ratio of isovanillin, harmine, and curcuminwas used, as compared with GZ17-6.02, namely ⅓ by weight of eachcomponent. This formulation was prepared by mixing the three GZ17-6.02components in 1 mL of ethanol to obtain a secondary stock solution. FIG.17A shows that the original stock solution (filled circles) and thesecondary stock solution (open circles) are bioequivalent in terms ofovarian cancer cell (A1847) kill rate. FIGS. 17B and 17C show the samegeneral effect for lung cancer cells (H358) and prostate cancer cells(22rv1).

Example 18

In this Example, a known ovarian cancer cell kill rate for GZ17-6.02 wasselected, and the amount of isovanillin in this dosage (0.78 mg/mL) wastested against GZ17-6.02. Additionally, sub-combinations ofisovanillin:curcumin and isovanillin:harmine were tested, where theconcentrations of the two ingredients were identical to those in theGZ17-6.02. Thus, the concentration of the isovanillin:curcumin productwas 0.78 mg/mL isovanillin and 0.13 mg/mL curcumin, and theconcentration of the isovanillin:harmine product was 0.78 mg/mLisovanillin and 0.099 mg/mL harmine. FIGS. 18A-18D illustrate theseresults with ovarian cancer cells (A1847), lung cancer cells (H358),prostate cancer cells (22rv1), and lymphoma cells (MO205). As isevident, the two-component products each exhibited greater kill rates ascompared with isovanillin alone.

Example 19

The procedures of Example 18 were followed, except that curcumin wastested alone versus GZ17-6.02, together with curcumin:isovanillin andcurcumin:harmine two-component products. In this test, the curcumin waspresent at a level of 0.78 mg/mL and the concentrations in thecurcumin:isovanillin product were 0.78 mg/mL curcumin and 3.59 mg/mLisovanillin, and in the curcumin:harmine product the concentrations were0.78 mg/mL curcumin and 0.59 mg/mL harmine. FIGS. 19A-19D illustrate theresults of this test, and confirm that the two-component products givebetter results than curcumin alone.

Example 20

The procedures of Example 18 were followed, except that harmine wastested alone versus GZ17-6.02, together with harmine:isovanillin andharmine:curcumin two-component products. In this test, the harmine waspresent at a level of 0.78 mg/mL and the concentrations in theharmine:isovanillin product were 0.78 mg/mL harmine and 6.09 mg/mLisovanillin, and in the harmine:curcumin product the concentrations were0.78 mg/mL curcumin and 1.03 mg/mL curcumin. FIGS. 20A-20D illustratethe results of this test, and confirm that the two-component productsgive better results than harmine alone.

Example 21

In this example, the individual components of GZ17-6.02 were testedagainst lymphoma cancer cells (MO205) at the component concentrationsfound in GZ17-6.02, at dosage rates of 12, 24, 48, and 96 μg/mL. Thetheoretical additive effect (black bar) of the 3 components was alsocalculated in each case and shown versus the actual test results foundusing GZ17-6.02. FIGS. 21A-21D illustrate the results of these tests,and confirm that, at the tested dosages, the GZ17-6.02 product had agreater effect than the individual components and the theoreticaladditive effect thereof, thus establishing synergistic effects. In thisregard, those skilled in the art understand that dosages of anticancerproducts actually administered to patients depend on a number offactors, including age, weight, sex, physical conditions, types ofcancers, and stages of cancers. By the same token, it should also beunderstood that at some dosage levels and/or specific concentrations ofcomponents, the results of FIGS. 21A-21D may not be duplicated. Thesesame considerations apply to the tests set forth in Examples 22 and 23below. In most cases, however, compositions giving therapeutic synergiesat the selected dosages are preferred.

Example 22

In this example, the individual components of GZ17-6.02 were testedagainst ovarian cancer cells (A1847) and breast cancer cells (du4475) atthe component concentrations found in GZ17-6.02, at dosage rates of 12and 24 μg/mL. The theoretical additive effect (black bar) of the 3components was also calculated in each case and shown versus the actualtest results found using GZ17-6.02. FIGS. 22A and 22B illustrate theresults of these ovarian cancer tests, and confirm that, at the testeddosages, the GZ17-6.02 product had a greater effect than the individualcomponents and the theoretical additive effect thereof. In an additionaltest at 48 μg/mL dosage rate, this effect was not observed. FIG. 22Csets forth the results using breast cancer cells.

Example 23

In this example, increasing concentrations of different isovanillinderivatives or metabolites were tested using lung cancer cells (H358),ovarian cancer cells (A1847), and prostate cancer cells (22rv1), todetermine the anticancer effects of the derivatives/metabolites, usingthe techniques of Example 1. Each of the derivatives/metabolites(vanillin, isovanillic acid, O-vanillin, and isovanillyl alcohol)demonstrated anticancer effects (see FIGS. 23A-23L).

Example 24

In this example, increasing concentrations of different harminederivatives or metabolites were tested using lung cancer cells (H358),ovarian cancer cells (A1847), and prostate cancer cells (22rv1), todetermine the anticancer effects of the derivatives/metabolites, usingthe techniques of Example 1. Each of the derivatives/metabolites(harmaline, tetrahydro-harmine, harmole hydrochloride, harmalolhydrochloride, and harmane) demonstrated anticancer effects (see FIGS.24A-24O).

Example 25

In this example, increasing concentrations of a curcumin derivative,bisdemethoxy curcumin, were tested using lung cancer cells (H358),ovarian cancer cells (A1847), and prostate cancer cells (22rv1), todetermine the anticancer effects of the derivative, using the techniquesof Example 1. The results confirmed the anticancer effects of thisderivative (see FIGS. 25A-25C).

Example 26

In this test, the preferred GZ17-6.02 product was stored at varioustemperatures over a two-month period, and then tested against ovariancancer cells (A1847). The results are set forth in

FIG. 26, which illustrates that at storage temperatures of −20° C. and4° C., the product maintained its potency.

Example 27

In this test, the GZ17-6.02 product was subjected to successivefreeze/thaw cycles. In each cycle, the product was frozen at −20° C.,followed by allowing the product temperature to equilibrate at roomtemperature. At the end of each cycle, the product was tested againstovarian cancer cells (A1847) using the methods of Example 1. A total of10 successive freeze/thaw cycles were performed on the same sample. FIG.27 illustrates that there was no significant change in the efficacy ofthe GZ17-6.02 against ovarian cancer cells.

Example 28

In this series of tests, GZ17-8.02 (which is identical with thepreferred GZ17-6.02 composition), and several different combinations ofcurcumin, harmine, and isovanillin derivatives were tested againstovarian cancer (A1847), lung cancer (H358), prostate cancer (22rv1),pancreatic cancer, and fibroblast cells (hgf-1), in order to confirmthat such derivative combinations were effective. In each case, theweight ratio of the isovanillin component:curcumin component:harminecomponent was 7.7:1:1.3, and the compositions were prepared as describedabove and the tests were carried out as described in Examples 1, 2, and6. The compositions are identified below, including the respectivegraphs giving the results of the tests:

-   -   GZ17-8.02 (FIG. 28A)    -   GZI7-8.03 contained vanillin, turmeric-derived curcumin, and        harmine (FIG. 28B)    -   GZI7-8.04 contained orthovanillin, turmeric-derived curcumin,        and harmine, and (FIG. 28C)    -   GZI7-8.05 contained isovanillyl alcohol, turmeric-derived        curcumin, and harmine (FIG. 28D)    -   GZI7-8.06 contained isovanillic acid, turmeric-derived curcumin,        and harmine (FIG. 28E)    -   GZI7-8.07 contained isovanillin, turmeric-derived curcumin, and        harmaline (FIG. 28F)    -   GZI7-8.08 contained isovanillin, turmeric-derived curcumin, and        harmane (FIG. 28G)    -   GZI7-8.09 contained isovanillin, 100% synthetic curcumin, and        harmine (FIG. 28H)    -   GZI7-8.10 contained isovanillin, bisdemethoxy curcumin, and        harmine (FIG. 28I) Examples 29-79

In each of the following examples, respective compositions havingdifferent combinations of isovanillin, curcumin, and harmine componentswere tested as set forth in Examples 1-6, against cancer cells todetermine the anti-cancer effectiveness thereof. The specific cancercell lines used in each case were those previously described. In allcases, the weight ratio of the isovanillin component:curcumincomponent:harmine component was 7.7:1:1.3, and the compositions wereprepared as described above in connection with the preparation ofGZ17-6.02. A separate graph is provided for each test, which identifiesthe cancer cells tested.

The following Table sets forth the Example numbers, compositiondesignation, components, and corresponding Figure numbers for thisseries of tests. Where “curcumin” is specified, this refers toturmeric-derived curcumin, and where “curcumin (syn)” is specified, thisrefers to essentially pure synthetically-derived curcumin.

Example Isovanillin Curcumin Harmine FIG. No. Composition ComponentComponent Component Nos. 29 GZ17-08.11 isovanillin bisdemethoxyharmaline 29 A-G curcumin 30 GZ17-08.12 isovanillin bisdemethoxy harmane30 A-G curcumin 31 GZ17-08.13 isovanillin bisdemethoxy harmalol 31 A-Gcurcumin 32 GZ17-08.14 isovanillin bisdemethoxy harmol 32 A-G curcumin33 GZ17-08.15 vanillin bisdemethoxy harmine 33 A-G curcumin 34GZ17-08.16 vanillin bisdemethoxy harmaline 34 A-G curcumin 35 GZ17-08.17vanillin bisdemethoxy harmane 35 A-G curcumin 36 GZ17-08.18 vanillinbisdemethoxy harmalol 36 A-G curcumin 37 GZ17-08.19 vanillinbisdemethoxy harmol 37 A-G curcumin 38 GZ17-08.20 isovanillin curcumin(syn) harmalol 38 A-G 39 GZ17-08.21 isovanillin curcumin (syn) harmol 39A-G 40 GZ17-08.22 vanillin curcumin (syn) harmaline 40 A-G 41 GZ17-08.23vanillin curcumin (syn) harmane 41 A-G 42 GZ17-08.24 vanillin curcumin(syn) harmalol 42 A-G 43 GZ17-08.25 vanillin curcumin (syn) harmol 43A-G 44 GZ17-08.26 orthovanillin bisdemethoxy harmine 44 A-G curcumin 45GZ17-08.27 orthovanillin bisdemethoxy harmaline 45 A-G curcumin 46GZ17-08.28 orthovanillin bisdemethoxy harmane 46 A-G curcumin 47GZ17-08.29 orthovanillin bisdemethoxy harmalol 47 A-G curcumin 48GZ17-08.30 orthovanillin bisdemethoxy harmol 48 A-G curcumin 49GZ17-08.31 isovanillyl bisdemethoxy harmine 49 A-D alcohol curcumin 50GZ17-08.32 isovanillyl bisdemethoxy harmaline 50 A-D alcohol curcumin 51GZ17-08.33 isovanillyl bisdemethoxy harmane 51 A-D alcohol curcumin 52GZ17-08.34 isovanillyl bisdemethoxy harmalol 52 A-D alcohol curcumin 53GZ17-08.35 isovanillyl bisdemethoxy harmol 53 A-D alcohol curcumin 54GZ17-08.36 orthovanillin curcumin (syn) harmaline 54 A-D 55 GZ17-08.37orthovanillin curcumin (syn) harmane 55 A-D 56 GZ17-08.38 orthovanillincurcumin (syn) harmalol 56 A-D 57 GZ17-08.39 orthovanillin curcumin(syn) harmol 57 A-D 58 GZ17-08.40 isovanillyl curcumin (syn) harmaline58 A-D alcohol 59 GZ17-08.41 isovanillyl curcumin (syn) harmane 59 A-Dalcohol 60 GZ17-08.42 isovanillyl curcumin (syn) harmalol 60 A-D alcohol61 GZ17-08.43 isovanillyl curcumin (syn) harmol 61 A-D alcohol 62GZ17-08.44 isovanillic acid bisdemethoxy harmine 62 A-D curcumin 63GZ17-08.45 isovanillic acid bisdemethoxy harmaline 63 A-D curcumin 64GZ17-08.46 isovanillic acid bisdemethoxy harmane 64 A-D curcumin 65GZ17-08.47 isovanillic acid bisdemethoxy harmalol 65 A-D curcumin 66GZ17-08.48 isovanillic acid bisdemethoxy harmol 66 A-D curcumin 67GZ17-08.49 isovanillic acid curcumin (syn) harmaline 67 A-D 68GZ17-08.50 isovanillic acid curcumin (syn) harmane 68 A-D 69 GZ17-08.51isovanillic acid curcumin (syn) harmalol 69 A-D 70 GZ17-08.52isovanillic acid curcumin (syn) harmol 70 A-D 71 GZ17-08.535-bromovanillin curcumin harmine 71 A-D 72 GZ17-08.54 2-bromo-3-curcumin harmine 72 A-D hydroxy-4- methoxy benzaldehyde 73 GZ17-08.552-iodo-3- curcumin harmine 73 A-D hydroxy-4- methoxy benzaldehyde 74GZ17-08.56 isovanillin (1E,4E)-1,5-bis(3,5- harmine 74 A-Ddimethoxyphenyl)-1,4- pentadien-3-one 75 GZ17-08.57 isovanillincardamonin harmine 75 A-D 76 GZ17-08.58 isovanillin2′-hydroxy-3,4,4′,5′- harmine 76 A-D tetramethoxychalcone 77 GZ17-08.59isovanillin 2,2′-dihydroxy-4′,6′- harmine 77 A-D dimethoxychalcone 78GZ17-08.60 isovanillin (1E,4E)-1,5-Bis(2- harmine 78 A-DHydroxyphenyl)-1,4- pentadien-3-one 79 GZ17-08.61 isovanillin curcumin1,2,3,4- 79 A-D tetrahy- droharmane- 3-carboxylic acid

These test results confirm that all of the compositions were effectiveanti-cancer agents.

Example 80

In this series of tests, the following compositions were prepared:

Isovanillin Curcumin Harmine Composition Component Component ComponentGZ17-10.00 orthovanillin curcumin harmaline GZ17-10.01 orthovanillin —harmaline GZ17-10.02 orthovanillin curcumin — GZ17-10.03 — curcuminharmaline GZ17-10.04 orthovanillin — — GZ17-10.05 — curcumin —GZ17-10.06 — — harmalineGZ17-10.00 is identical with the above-described GZ17-6.02, having theidentical amounts of the components and method of preparation. Thetwo-component compositions have the same relative ratios, namely 7.7:1.3for GZ17-10.01, 7.7:1 for GZ17-10.02, and 1:1.3 for GZ17-10.03. Thesingle-component compositions contain the same amount of component asused in the three- and two-component compositions.

The respective compositions of this Example were tested againstdifferent cancer cell lines, as previously identified, using the methodsof Examples 1-6, as set forth in the accompanying relevant graphs,80A-80R. Six of these (FIGS. 80G, H, L, M, Q, and R) are comparative bargraphs setting forth the theoretical additive effect of the components,and the results obtained using the complete formulations, therebydemonstrating the synergistic effects of the compositions of theinvention.

These test results confirm that all of the compositions were effectiveanti-cancer agents.

Example 81

In this series of tests, the following compositions were prepared:

Isovanillin Component Harmine Component Composition (ratio component =7.7) (ratio component = 1.3) GZ17-08.512 isovanillin harmine GZ17-08.513isovanillin harmaline GZ17-08.514 isovanillin harmane GZ17-08.515isovanillin harmalol GZ17-08.516 isovanillin harmol GZ17-08.517 vanillinharmine GZ17-08.518 vanillin harmaline GZ17-08.519 vanillin harmaneGZ17-08.520 vanillin harmalol GZ17-08.521 vanillin harmol

The respective compositions of this Example were tested againstdifferent cancer cell lines, as previously identified, using the methodsof Examples 1-6, as set forth in the accompanying relevant graphs,81A-81DD.

These test results confirm that all of the compositions were effectiveanti-cancer agents.

Example 82

In each of the following examples, respective compositions havingdifferent combinations of isovanillin, curcumin, and harmine componentswere tested as set forth in Examples 1-3, against lung cancer, lymphoma,and leukemia cells to determine the anti-cancer effectiveness thereof.The compositions were prepared by individually mixing the listedcomponents in 3 mL of ethanol followed by adding the so-mixed componentstogether to form the resultant compositions. A separate graph isprovided for each test, which identifies the cancer cells tested.Specifically, FIGS. 82-1 through 82-41 set forth the results of the lungcancer (LC) tests using the compositions, FIGS. 82-42 through 82-82 givethe results for the lymphoma (LY) tests, and FIGS. 82-83 through 82-123give the results of the leukemia (LK) tests.

The first Table below sets forth the identities of the isovanillincomponents i01-i15, the harmine components h01-h13, and the curcumincomponents c01-c13, used in these tests. The second Table below setsforth the multiple-component compositions tested, GZ08.065-GZ08.105, forlung cancer, lymphoma, and leukemia cells and the respective Figurenumbers associated with each such composition and cell line. Therecitation “curcumin (syn)” refers to essentially puresynthetically-derived curcumin.

All of the second Table compositions exhibited anti-cancer activityagainst lung cancer, lymphoma, and leukemia cells. In addition, FIGS.82-124 through 82-167 are comparative bar graphs confirming thesynergism found in exemplary compositions of this Example, at variousconcentration levels, against lung cancer, lymphoma, and leukemia cells.

Code Compound CAS# MW i01 o-anisaldehyde 135-02-4 136.15 i02 isovanillinoxime 51673-94-0 167.166 i03 ethyl vanillin 121-32-4 166.18 i04 vanillinisobutyrate 20665-85-4 222.24 i05 veratraldehyde 120-14-9 166.18 i065-nitrovanillin 6635-20-7 197.14 i07 vanillin acetate 881-68-5 194.18i08 3-benzyloxy-4-methoxybenzaldehyde 6346-05-0 242.27 i093-hydroxy-5-methoxybenzaldehyde 672-13-9 152.15 i10 methyl isovanillate6702-50-7 182.18 i11 acetovanillone (apocynin) 498-02-2 166.17 i122-hydroxy-4-methoxybenzaldehyde 673-22-3 152.15 i13 trans-ferulic acid537-98-4 194.18 i14 3-hydroxy-4-methoxycinnamic acid 537-73-5 194.18 i15caffeic acid 331-39-5 180.16 h01 norharmane 244-63-3 168.19 h026-methoxyharmalan 3589-73-9 214.26 h03 bromo harmine na 372.06 h042-methyl harmine 21236-68-0 227.28 h05 4,9-dihydro-3H-beta-carbolin-1-ylna 200.24 methyl ether h06 1-(4-nitrophenyl)-2,3,4,9-tetrahydro-3380-77-6 329.79 1H-beta-carboline hydrochloride h071,2,3,4-tetrahydro-9H-pyrido[3,4- 16502-01-5 172.23 b]indole {THbC} h081,2,3,4-tetrahydro-beta-carboline-1- na 216.24 carboxylic acid h096-Methoxy-1,2,3,4-tetrahydro-9H- 20315-68-8 202.25 pyrido[3,4-b]indoleh10 3-hydroxymethyl-b-carboline 65474-79-5 198.22 h112,3,4,5-tetrahydro-8-methoxy-1H- na 202.25 pyrido[4,3-b]indole h126-Methoxy-1,2,3,4-tetrahydro-9H- 17952-63-5 246.26pyrido[3,4-b]indole-1-carboxylic acid h13 ethylb-carboline-3-carboxylate 74214-62-3 240.26 c01 tetrahydro curcumin36062-04-1 372.41 c02 demethyl curcumin 149732-51-4 354.35 c03 demethoxycurcumin 22608-11-3 338.35 c04 1,3-diphenyl-2-propanone 102-04-5 210.27c05 caffeic acid phenethyl ester 104594-70-9 284.31 c06(1E,4E)-1,5-bis[3,5-bis(methoxy- 917813-62-8 474.51methoxy)phenyl]-1,4-pentadiene-3-one c071,7-di(1-naphthyl)-2,6-heptanedione na 380.49 c08trans,trans-1,5-Bis[4-(trifluoro- 103836-71-1 370.29methyl)phenyl]-1,4-pentadien-3-one c09 1,5-dibenzoylpentane 28861-22-5280.36 c10 (2E,5E)-2,5- 895-80-7 260.33 dibenzylidenecyclopentanone c112,6-bis(4-fluorobenzal)cyclohexanone 62085-74-9 310.34 c12(1E,4E)-1,5-bis(4-fluorophenyl)-1,4- na 354.4 pentadien-3-one c13 FLLL3152328-97-9 424.49

Isovanillin Harmine Curcumin Component Component Component LC LY LK (176mg/3 mL (30 mg/3 mL (23 mg/3 mL Total Total FIG. FIG. FIG. Comp. EtOH)EtOH) EtOH) (mg/mL) (mM) Nos. Nos. Nos. 08.065 i01 harmine curcumin(syn) 76.3 498.8 82-1 82-42 82-83 08.066 i02 harmine curcumin (syn) 76.3418.8 82-2 82-43 82-84 08.067 I03 harmine curcumin (syn) 76.3 420.9 82-382-44 82-85 08.068 i04 harmine curcumin (syn) 76.3 331.9 82-4 82-4582-86 08.069 i05 harmine curcumin (syn) 76.3 420.9 82-5 82-46 82-8708.070 i06 harmine curcumin (syn) 76.3 365.5 82-6 82-47 82-88 08.071 i07harmine curcumin (syn) 76.3 370.0 82-7 82-48 82-89 08.072 i08 harminecurcumin (syn) 76.3 310.1 82-8 82-49 82-90 08.073 i09 harmine curcumin(syn) 76.3 453.5 82-9 82-50 82-91 08.074 i10 harmine curcumin (syn) 76.3389.9 82-10 82-51 82-92 08.075 i11 harmine curcumin (syn) 76.3 421.082-11 82-52 82-93 08.076 i12 harmine curcumin (syn) 76.3 453.5 82-1282-53 82-94 08.077 i13 harmine curcumin (syn) 76.3 370.0 82-13 82-5482-95 08.078 i14 harmine curcumin (syn) 76.3 370.0 82-14 82-55 82-9608.079 i15 harmine curcumin (syn) 76.3 393.5 82-15 82-56 82-97 08.080isovanillin h01 curcumin (syn) 76.3 465.9 82-16 82-57 82-98 08.081isovanillin h02 curcumin (syn) 76.3 453.1 82-17 82-58 82-99 08.082isovanillin h03 curcumin (syn) 76.3 433.3 82-18 82-59 82-100 08.083isovanillin h04 curcumin (syn) 76.3 450.4 82-19 82-60 82-101 08.084isovanillin h05 curcumin (syn) 76.3 456.3 82-20 82-61 82-102 08.085isovanillin h06 curcumin (syn) 76.3 436.7 82-21 82-62 82-103 08.086isovanillin h07 curcumin (syn) 76.3 464.5 82-22 82-63 82-104 08.087isovanillin h08 curcumin (syn) 76.3 452.6 82-23 82-64 82-105 08.088isovanillin h09 curcumin (syn) 76.3 455.8 82-24 82-65 82-106 08.089isovanillin h10 curcumin (syn) 76.3 456.8 82-25 82-66 82-107 08.090isovanillin h11 curcumin (syn) 76.3 455.8 82-26 82-67 82-108 08.091isovanillin h12 curcumin (syn) 76.3 447.0 82-27 82-68 82-109 08.092isovanillin h13 curcumin (syn) 76.3 448.0 82-28 82-69 82-110 08.093isovanillin harmine c01 76.3 453.3 82-29 82-70 82-111 08.094 isovanillinharmine c02 76.3 454.3 82-30 82-71 82-112 08.095 isovanillin harmine c0376.3 455.4 82-31 82-72 82-113 08.096 isovanillin harmine c04 76.3 469.282-32 82-73 82-114 08.097 isovanillin harmine c05 76.3 459.7 82-33 82-7482-115 08.098 isovanillin harmine c06 76.3 448.9 82-34 82-75 82-11608.099 isovanillin harmine c07 76.3 452.8 82-35 82-76 82-117 08.100isovanillin harmine c08 76.3 453.4 82-36 82-77 82-118 08.101 isovanillinharmine c09 76.3 460.0 82-37 82-78 82-119 08.102 isovanillin harmine c1076.3 462.1 82-38 82-79 82-120 08.103 isovanillin harmine c11 76.3 457.482-39 82-80 82-121 08.104 isovanillin harmine c12 76.3 454.3 82-40 82-8182-122 08.105 isovanillin harmine c13 76.3 450.8 82-41 82-82 82-123

All of the foregoing tests confirmed significant anti-cancer activityagainst lung cancer cells, H358, lymphoma cells, MO205, and leukemiacells, jurkat E6-1. Accordingly, all combinations of i01-i15, h01-h13,and c01-c13, whether three-component or two-component, would exhibitsuch anti-cancer activity. Moreover, as shown above, the combinations ofharmine and curcumin with i01-15, isovanillin and curcumin with h01-h13,and isovanillin and harmine with c01-c13 all have useful anti-canceractivity.

Moreover, as noted previously, the compounds i01415, h01-h13, andc01-c13 may be individually and independently in the form of thecorresponding esters, metal complexes, pharmaceutically acceptablesalts, and mixtures thereof. That is, e.g., a given curcumin componentmay be in the form of an ester, complex, or salt, independently of theform of the remaining components of the composition.

Example 83

In another series of tests, compositions were prepared containing: (1)orthovanillin+curcumin+harmaline; and (2) single-component compositionscontaining orthovanillin, curcumin, and harmaline, respectively. Thethree-component composition had a weight ratio oforthovanillin:curcumin:harmaline of 771:130.3:98.7, and thesingle-component compositions contained varying amounts oforthovanillin, curcumin, and harmaline.

These compositions were prepared by mixing together quantities of solidsynthetic orthovanillin (99% by weight purity), synthetic harmaline (92%by weight purity), and a commercial turmeric product (ResCu) in ethanol,and allowing the mixtures to react for a period of 24 hours.

These compositions were tested against lymphoma (MO205), leukemia(jurkat E6-1), and breast cancer (du4475) cell lines, using the assaysdescribed in Examples 1-3.

The results of the lymphoma tests are set forth in FIGS. 83A-83D. Thesetests confirmed that the combination of orthovanillin+curcumin+harmaline(FIG. 83D) exhibited synergistic results as compared with the individualtests of orthovanillin, curcumin, and harmaline (FIGS. 83A-83C).

In like manner, the leukemia tests (FIGS. 83E-83H) confirmed that thecombination of orthovanillin+curcumin+harmaline (FIG. 83H) exhibitedsynergistic results as compared with the individual tests oforthovanillin, curcumin, and harmaline (FIGS. 83E-83G).

To a similar effect, the breast cancer tests (FIGS. 83I-83L) confirmedthat the combination of orthovanillin+curcumin+harmaline (FIG. 83L)exhibited synergistic results as compared with the individual tests oforthovanillin, curcumin, and harmaline (FIGS. 83I-83K).

Example 84

In this Example, GZ17-6.02 was tested with pancreatic cancer cells(S2-007) to determine whether it had an effect on cancer stem cellproteins using the techniques described in Examples 1-3, and it wasfound that the treatment decreased the number and size of the pancreaticcancer spheres at both the IC₅₀ concentration and a concentration ofhalf that amount (see FIG. 84A). In additional tests (FIG. 84B), treatedcells exhibited a dose-dependent decline in doublecortin calmodulin-likekinase 1 (Dclk-1), a microtubule-associated protein that serves as amarker for intestinal and pancreatic cancer stem cells (Mwangi, S. M. etal. “DCAMKL-1: a new horizon for pancreatic progenitor identification.”Am J Phys-Gastro Liver Physiol 299 (2010): G301-302), and alsodifferentiates between tumor stem cells and normal stem cells(Nakanishi, Y. et al. “Dclk1 distinguishes between tumor and normal stemcells in the intestine.” Nature Gen 45 (2013):98-103). Epithelial celladhesion molecule (EpCAM) is another cancer stem cell marker and it alsodecreased in level with GZ17-6.02 exposure. Likewise, theleucine-rich-repeat containing G-protein-coupled receptor 5 (LGR5) isanother cancer stem cell marker and its levels decreased with theGZ17-6.02 treatment (FIG. 84B).

When tumors from treated mice were analyzed, the tumors also showed aclear reduction in EPCAM, DCLK1, LGR5, and SOX9 (FIG. 84C). GZ17-6.02 isthus seen to block pancreatic spheroid formation and cancer stem cellmarker expression, both in cancer cells grown and treated in vitro andin tumor samples from mice treated with GZ17-6.02.

GZ17-6.02 significantly reduced tumorigenesis both with in vitro and invivo pancreatic cancer cell models, partially via a route that inhibitsthe presence of cancer stem cells. In this regard, it appears thatGZ17-6.02 has several different mechanisms of action resulting indecreased tumor growth and inhibition of metastases. FIG. 84Dillustrates a possible pathway for the action of GZ17-6.02 on bothcancer stem cells (CSC) and on the other cancer cells within the tumor.With respect to the cancer stem cells, the GZ17-6.02 decreased thecommon biomarkers of cancer stem cells (Dclk1, LGR5, EpCam, and Sox9).It apparently did this by acting through the Sonic Hedgehog pathway(FIG. 84D, SHH). Simultaneously, GZ17-6.02 blocked the bulk tumor cellsin two ways by directly inducing apoptosis via the caspase pathway, asmeasured by the BAX/Bcl2 ratio, and through the EGFR pathway. All threeof these mechanisms led to reduced tumor size, inhibition of metastasisand decline in the presence of cancer stem cells. Thus, the compositionsof the invention are effective for killing or inhibiting the growth ofcancer stem cells.

Example 85

The purpose of this study was to determine the effect of GZ17-6.02 onthe regression of growth and the inhibition of metastases in apancreatic cancer mouse model. Twelve immunocompromised (nude) mice weretreated injected with MiaPaCa-2 cells (a pancreatic cancer cell line)that had been genetically transformed to express TdTomato and luciferinfor in vivo tumor imaging. After tumors reached a measurable size(approximately 2 weeks), half of the mice were randomly assigned to theexperimental group and the other half to the control group.

The experimental group was treated with GZ17-6.02 at a daily dose of 100mg/kg body weight. The drug administration was oral, dissolved inpeptamen. Control mice were fed the vehicle (peptamen) alone. Once aweek mice were lightly anesthetized in order to use IVIS imaging todetect the tumor sizes. On day 5, there was no statistical difference inthe amount of fluorescence measured from the control (placebo-fed) andGZ17-6.02-fed mice. However, by day 20, there was a statisticallysignificant decrease in the amount of fluorescence, indicating adecreased total tumor burden, in the mice fed GZ17-6.02, as comparedwith the control. In addition, the placebo-treated mice had dramaticlevels of peritoneal ascites that were not present in the GZ17-6.02group.

At the end of 21 days of treatment with GZ17-6.02 or the placebo, themice were sacrificed and tumor samples removed from the pancreas, liver,and lungs. The primary pancreatic tumors were weighed. The placebo-fedanimals had pancreatic tumors that averaged 2.4 times statisticallylarger than the GZ17-6.02 fed mice.

Example 86

A 63-year-old male patient having a Prostate-Specific Antigen (PSA)count of 8.2 began a treatment regimen using a dosage form of GZ17-6.02.Specifically, solid GZ17-6.02 prepared by evaporating the ethanol fromthe previously described GZ17-6.02 agent was dispersed in water in equalweight amounts, e.g., 5 grams solid GZ17-6.02 in 5 grams water. Thisdosage form was taken three times daily for six weeks, with each dosebeing four fluid ounces of the 50%:50% dispersion. At the end of sixweeks, the patient's PSA count had dropped to 2.1, and all prostate andurological tests were normal. The treating physician had no explanationfor the decline in PSA. The patient experienced no observable adversereactions to the treatment with GZ17-6.02.

Although not wishing to be bound by any theory of operation, theinventors believe that the therapeutic agents of the inventionameliorate a number of conditions or illnesses, and especially reduceand/or eliminate cancer and/or the symptoms thereof by augmenting orstimulating the patients' immune systems. In this sense, the inventionis believed to be a form of biological therapy. As such, it isconsidered that the invention is applicable to virtually all cancers,such as the following: Acute Lymphoblastic Leukemia, Adult; AcuteLymphoblastic Leukemia, Childhood; Acute Myeloid Leukemia, Adult; AcuteMyeloid Leukemia, Childhood; Adrenocortical Carcinoma; AdrenocorticalCarcinoma, Childhood; Adolescents, Cancer in; AIDS-Related Cancers;AIDS-Related Lymphoma; Anal Cancer; Appendix Cancer; Astrocytomas,Childhood; Atypical Teratoid/Rhabdoid Tumor, Childhood, Central NervousSystem; Basal Cell Carcinoma; Bile Duct Cancer, Extrahepatic; BladderCancer; Bladder Cancer, Childhood; Bone Cancer, Osteosarcoma andMalignant Fibrous Histiocytoma; Brain Stem Glioma, Childhood; BrainTumor, Adult; Brain Tumor, Brain Stem Glioma, Childhood; Brain Tumor,Central Nervous System Atypical Teratoid/Rhabdoid Tumor, Childhood;Brain Tumor, Central Nervous System Embryonal Tumors, Childhood; BrainTumor, Astrocytomas, Childhood; Brain Tumor, Craniopharyngioma,Childhood; Brain Tumor, Ependymoblastoma, Childhood; Brain Tumor,Ependymoma, Childhood; Brain Tumor, Medulloblastoma, Childhood; BrainTumor, Medulloepithelioma, Childhood; Brain Tumor, Pineal ParenchymalTumors of Intermediate Differentiation, Childhood; Brain Tumor,Supratentorial Primitive Neuroectodermal Tumors and Pineoblastoma,Childhood; Brain and Spinal Cord Tumors, Childhood (Other); BreastCancer; Breast Cancer and Pregnancy; Breast Cancer, Childhood; BreastCancer, Male; Bronchial Tumors, Childhood; Burkitt Lymphoma; CarcinoidTumor, Childhood; Carcinoid Tumor, Gastrointestinal; Carcinoma ofUnknown Primary; Central Nervous System Atypical Teratoid/RhabdoidTumor, Childhood; Central Nervous System Embryonal Tumors, Childhood;Central Nervous System (CNS) Lymphoma, Primary; Cervical Cancer;Cervical Cancer, Childhood; Childhood Cancers; Chordoma, Childhood;Chronic Lymphocytic Leukemia; Chronic Myelogenous Leukemia; ChronicMyeloproliferative Disorders; Colon Cancer; Colorectal Cancer,Childhood; Craniopharyngioma, Childhood; Cutaneous T-Cell Lymphoma;Embryonal Tumors, Central Nervous System, Childhood; Endometrial Cancer;Ependymoblastoma, Childhood; Ependymoma, Childhood; Esophageal Cancer;Esophageal Cancer, Childhood; Esthesioneuroblastoma, Childhood; EwingSarcoma Family of Tumors; Extracranial Germ Cell Tumor, Childhood;Extragonadal Germ Cell Tumor; Extrahepatic Bile Duct Cancer; Eye Cancer,Intraocular Melanoma; Eye Cancer, Retinoblastoma; Gallbladder Cancer;Gastric (Stomach) Cancer; Gastric (Stomach) Cancer, Childhood;Gastrointestinal Carcinoid Tumor; Gastrointestinal Stromal Tumor (GIST);Gastrointestinal Stromal Cell Tumor, Childhood; Germ Cell Tumor,Extracranial, Childhood; Germ Cell Tumor, Extragonadal; Germ Cell Tumor,Ovarian; Gestational Trophoblastic Tumor; Glioma, Adult; Glioma,Childhood Brain Stem; Hairy Cell Leukemia; Head and Neck Cancer; HeartCancer, Childhood; Hepatocellular (Liver) Cancer, Adult (Primary);Hepatocellular (Liver) Cancer, Childhood (Primary); Histiocytosis,Langerhans Cell; Hodgkin Lymphoma, Adult; Hodgkin Lymphoma, Childhood;Hypopharyngeal Cancer; Intraocular Melanoma; Islet Cell Tumors(Endocrine Pancreas); Kaposi Sarcoma; Kidney (Renal Cell) Cancer; KidneyCancer, Childhood; Langerhans Cell Histiocytosis; Laryngeal Cancer;Laryngeal Cancer, Childhood; Leukemia, Acute Lymphoblastic, Adult;Leukemia, Acute Lymphoblastic, Childhood; Leukemia, Acute Myeloid,Adult; Leukemia, Acute Myeloid, Childhood; Leukemia, ChronicLymphocytic; Leukemia, Chronic Myelogenous; Leukemia, Hairy Cell; Lipand Oral Cavity Cancer; Liver Cancer, Adult (Primary); Liver Cancer,Childhood (Primary); Lung Cancer, Non-Small Cell; Lung Cancer, SmallCell; Lymphoma, AIDS-Related; Lymphoma, Burkitt; Lymphoma, CutaneousT-Cell; Lymphoma, Hodgkin, Adult; Lymphoma, Hodgkin, Childhood;Lymphoma, Non-Hodgkin, Adult; Lymphoma, Non-Hodgkin, Childhood;Lymphoma, Primary Central Nervous System (CNS); Macroglobulinemia,Waldenström; Malignant Fibrous Histiocytoma of Bone and Osteosarcoma;Medulloblastoma, Childhood; Medulloepithelioma, Childhood; Melanoma;Melanoma, Intraocular (Eye); Merkel Cell Carcinoma; Mesothelioma, AdultMalignant; Mesothelioma, Childhood; Metastatic Squamous Neck Cancer withOccult Primary; Mouth Cancer; Multiple Endocrine Neoplasia Syndromes,Childhood; Multiple Myeloma/Plasma Cell Neoplasm; Mycosis Fungoides;Myelodysplastic Syndromes; Myelodysplastic/Myeloproliferative Neoplasms;Myelogenous Leukemia, Chronic; Myeloid Leukemia, Adult Acute; MyeloidLeukemia, Childhood Acute; Myeloma, Multiple; MyeloproliferativeDisorders, Chronic; Nasal Cavity and Paranasal Sinus Cancer;Nasopharyngeal Cancer; Nasopharyngeal Cancer, Childhood; Neuroblastoma;Non-Hodgkin Lymphoma, Adult; Non-Hodgkin Lymphoma, Childhood; Non-SmallCell Lung Cancer; Oral Cancer, Childhood; Oral Cavity Cancer, Lip and;Oropharyngeal Cancer; Osteosarcoma and Malignant Fibrous Histiocytoma ofBone; Ovarian Cancer, Childhood; Ovarian Epithelial Cancer; Ovarian GermCell Tumor; Ovarian Low Malignant Potential Tumor; Pancreatic Cancer;Pancreatic Cancer, Childhood; Pancreatic Cancer, Islet Cell Tumors;Papillomatosis, Childhood; Paranasal Sinus and Nasal Cavity Cancer;Parathyroid Cancer; Penile Cancer; Pharyngeal Cancer; Pineal ParenchymalTumors of Intermediate Differentiation, Childhood; Pineoblastoma andSupratentorial Primitive Neuroectodermal Tumors, Childhood; PituitaryTumor; Plasma Cell Neoplasm/Multiple Myeloma; Pleuropulmonary Blastoma,Childhood; Pregnancy and Breast Cancer; Primary Central Nervous System(CNS) Lymphoma; Prostate Cancer; Rectal Cancer; Renal Cell (Kidney)Cancer; Renal Pelvis and Ureter, Transitional Cell Cancer; RespiratoryTract Cancer with Chromosome 15 Changes; Retinoblastoma;Rhabdomyosarcoma, Childhood; Salivary Gland Cancer; Salivary GlandCancer, Childhood; Sarcoma, Ewing Sarcoma Family of Tumors; Sarcoma,Kaposi; Sarcoma, Soft Tissue, Adult; Sarcoma, Soft Tissue, Childhood;Sarcoma, Uterine; Sézary Syndrome; Skin Cancer (Nonmelanoma); SkinCancer, Childhood; Skin Cancer (Melanoma); Skin Carcinoma, Merkel Cell;Small Cell Lung Cancer; Small Intestine Cancer; Soft Tissue Sarcoma,Adult; Soft Tissue Sarcoma, Childhood; Squamous Cell Carcinoma; SquamousNeck Cancer with Occult Primary, Metastatic; Stomach (Gastric) Cancer;Stomach (Gastric) Cancer, Childhood; Supratentorial PrimitiveNeuroectodermal Tumors, Childhood; T-Cell Lymphoma, Cutaneous;Testicular Cancer; Testicular Cancer, Childhood; Throat Cancer; Thymomaand Thymic Carcinoma; Thymoma and Thymic Carcinoma, Childhood; ThyroidCancer; Thyroid Cancer, Childhood; Transitional Cell Cancer of the RenalPelvis and Ureter; Trophoblastic Tumor, Gestational; Unknown PrimarySite, Carcinoma of, Adult; Unknown Primary Site, Cancer of, Childhood;Unusual Cancers of Childhood; Ureter and Renal Pelvis, Transitional CellCancer; Urethral Cancer; Uterine Cancer, Endometrial; Uterine Sarcoma;Vaginal Cancer; Vaginal Cancer, Childhood; Vulvar Cancer; WaldenströmMacroglobulinemia; Wilms Tumor; Women's Cancers.

We claim:
 1. A therapeutic composition comprising a quantity of aharmine component selected from the group consisting of harmine, and theesters, metal complexes, and pharmaceutically acceptable salts thereof,with a quantity of a isovanillin component selected from the groupconsisting of vanillin, isovanillin, and mixtures thereof, and theesters, metal complexes, and pharmaceutically acceptable salts of theforegoing, said harmine component and said isovanillin component beingselected and present in amounts to provide anti-cancer synergy for thecomposition.
 2. The composition of claim 1, said isovanillin componentcomprising isovanillin.
 3. The composition of claim 1, wherein saidisovanillin component is present at a level greater than that of saidharmine component.
 4. The composition of claim 1, the weight ratio ofsaid isovanillin component to said harmine component ranging from about0.5:1 to 15:1.
 5. The composition of claim 1, said isovanillin componentbeing present of from about 25-95% by weight, and said harmine componentbeing present at a level of from about 5-75% by weight, based upon thetotal weight of the composition taken as 100% by weight.
 6. Thecomposition of claim 1, said composition exhibiting anti-cancer activityagainst ovarian, lung, prostate, lymphoma, leukemia, and head and neckcancer cells.
 7. The composition of claim 6, said composition exhibitinganti-cancer synergy against lung cancer cells, H358, lymphoma cells,M0205, and leukemia cells, Jurkat-E6-1.
 8. The composition of claim 1,said composition being in the form of liquid mixtures, gels,suspensions, solutions, solids, capsules, pills, or tablets.
 9. A methodof making a therapeutic composition in accordance with claim 1comprising the steps of mixing together a quantity of said harminecomponent, and a quantity of said isovanillin component.
 10. The methodof claim 9, said isovanillin component comprising isovanillin.
 11. Themethod of claim 9, wherein said isovanillin component is present at alevel greater than that of said harmine component.
 12. The method ofclaim 9, the weight ratio of said isovanillin component to said harminecomponent ranging from about 0.5:1 to 15:1.
 13. The method of claim 9,said isovanillin component being present of from about 25-95% by weight,and said harmine component being present at a level of from about 5-75%by weight, based upon the total weight of the composition taken as 100%by weight.
 14. The method of claim 9, said composition exhibitinganti-cancer activity against ovarian, lung, prostate, lymphoma,leukemia, and head and neck cancer cells.
 15. The method of claim 14,said composition exhibiting anti-cancer synergy against lung cancercells, H358, lymphoma cells, M0205, and leukemia cells, Jurkat-E6-1. 16.The method of claim 9, said composition being in the form of liquidmixtures, gels, suspensions, solutions, solids, capsules, pills, ortablets.